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G Cornelis

Publications and source records attributed to G Cornelis.

At least 37 records · Page 2Linked to original sources
PubMed1986

Tn951 derivatives designed for high-frequency plasmid-specific transposition and deletion mutagenesis.

We describe the construction of a system allowing high-frequency transposition and deletion mutagenesis with class-II transposons containing a kanamycin or a chloramphenicol-resistance marker. The system utilizes the transposition function of Tn3 and the resolution function of Tn951/Tn2501 which leads to an uncoupling of the resolution and repression functions. It consists of defective transposons inserted into conjugative, replication thermosensitive plasmids. The properties of the system are: easily selectable resistance markers, high transposition frequencies onto plasmids, low transposition frequencies onto the host chromosome, placement of the tnpA gene outside the transposons so that "second-generation" transposition does not occur, possibility to transpose the whole system onto other plasmid vectors with different selection strategies, consecutive use of two transposons for deletion mutagenesis and restriction mapping.

Chromosome Deletion↗
PubMed1986

Genetic analysis of the plasmid region controlling virulence in Yersinia enterocolitica 0:9 by Mini-Mu insertions and lac gene fusions.

Enteropathogenic strains of Yersinia enterocolitica harbor a virulence plasmid (pYVe plasmid) of 70 kilobases (kb) which specifies, at 37 degrees C, a calcium requirement for growth, autoagglutinability, resistance to the bactericidal activity of human serum, and the expression of outer membrane proteins (OMP). Some mutations suppress the calcium requirement for growth while others make the bacteria unable to grow at 37 degrees C, even in the presence of calcium. To analyse the genes involved in these latter phenotypes, the plasmid of a serogroup 0:9 strain was subjected to transposon mutagenesis with a Mini-Mu (Kan, lac) element. The mapping of 15 insertions and the analysis of transcription showed that at least four transcription units, spanning 22 kb, are involved in the phenomenon of calcium dependence. Mutations in two divergent units (virA and virB) suppressed the requirement for calcium at 37 degrees C. When insertions occurred in the other units (virC and virD), the Y. enterocolitica host became thermosensitive for growth. VirA, B and C mutants did no longer express the pYVe dependent OMPs. VirD mutants expressed and released these proteins save two of mol. wt 37,400 and 40,800 daltons. Transcription of the lac genes in the four groups of mutants was dependent on temperature.

Cloning, Molecular↗
PubMed1986

Comparison of serogrouping and polyacrylamide gel electrophoresis for typing Clostridium difficile.

A typing scheme for Clostridium difficile based on slide agglutination with rabbit antisera was previously described. It allows the differentiation of 10 serogroups designated A, B, C, D, F, G, H, I, K, and X. We studied the correlation between serogrouping and polyacrylamide gel electrophoresis (PAGE) of whole-cell proteins. A total of 202 isolates from different sources were analyzed by PAGE after ultrasonic disintegration of cells from an 18-h liquid culture and treatment with sodium dodecyl sulfate and 2-mercaptoethanol. A total of 21 different patterns were observed. The reference strains from the 10 serogroups showed different profiles. For each serogroup except A, the patterns obtained with the clinical isolates were identical to the patterns obtained with the reference strains. For the 48 strains belonging to serogroup A, 12 different profiles were observed. Five of these involved strains isolated from patients with antibiotic-associated diarrhea. Typing by sodium dodecyl sulfate-PAGE thus correlates with serogrouping. In addition, it allows discrimination within the heterogeneous serogroup A.

Agglutination Tests↗
PubMed1985

A simple adult-mouse test for tissue invasiveness in Yersinia enterocolitica strains of low experimental virulence.

The virulence of Yersinia enterocolitica depends on the presence of a 70-kilobase plasmid, called the Vwa plasmid. This situation is particularly favourable for studies of the mechanism of pathogenicity, but these are hindered by the lack of a suitable animal test to monitor the virulence of the human-pathogenic strains isolated outside the USA which belong to serogroups O:3, O:9 and O:5,27. We observed that, after oral administration to the mouse, the Vwa-positive strains of these serogroups produce a discrete systemic infection while the Vwa-negative strains do not. We present here a simple mouse-virulence test based on this observation.

Animals↗
PubMed1985

Genetic analysis of virulence plasmid from a serogroup 9 Yersinia enterocolitica strain: role of outer membrane protein P1 in resistance to human serum and autoagglutination.

Enteropathogenic strains of Yersinia enterocolitica harbor a virulence plasmid (70 kilobases) which specifies, at 37 degrees C, a calcium requirement for growth, autoagglutinability, resistance to the bactericidal activity of human serum, and the expression of some outer membrane proteins (OMPs). To map the genes encoding these properties, the virulence plasmid of a serogroup 9 strain (W22708) was subjected to transposon mutagenesis. A set of 68 independent mutations was obtained in Escherichia coli by transposon Tn813 (a tnpR mutant of Tn21)-mediated cointegration with the self-transmissible R388 plasmid. The resulting cointegrates were introduced and studied in Y. enterocolitica W22708. One mutant lost the calcium dependence property. Two other mutants presented a peculiar phenotype: they grew poorly at 37 degrees C, especially in the presence of calcium. Lastly, two mutants were affected in the properties of autoagglutination and resistance to human serum. Analysis of the OMP pattern of these two mutants revealed the absence of the largest OMP, called P1 (I. Bölin, and H. Wolf-Watz, Infect. Immun. 43:72-78, 1984). Complementation of one of these mutations with the cloned structural gene of OMP P1 restored the wild-type phenotype. However, OMP P1 was not sufficient by itself to specify the serum resistance property and a rapid autoagglutination of the host.

Agglutination↗
PubMed1984

A restriction map of virulence plasmid pVYE439-80 from a serogroup 9 Yersinia enterocolitica strain.

A restriction map of the virulence plasmid pVYE439-80, isolated from Yersinia enterocolitica 439-80 (serogroup 9) was constructed for EcoRI, BamHI, SstII, and SmaI. The mapping was done after cloning of about two-thirds of the plasmid in Escherichia coli. The restriction pattern was compared to those obtained with plasmids isolated from Y. enterocolitica strains of serogroups 1, 3, and 5b. The restriction sites are particularly conserved in a region of about 25 kb. This region contains fragments that are also conserved in serogroup 8 strains [J. Heeseman, C. Keller, R. Morawa, N. Schmidt, H. J. Siemens, and R. Lauf (1983) J. Infect. Dis. 147, 107-115] and that were shown, in strains from this serogroup, to encode calcium dependency [D. A. Portnoy, H. Wolf-Watz, I. Bolin, A. B. Beeder, and S. Falkow, (1984) Infect. Immun. 43, 108-114].

Base Sequence↗
PubMed1984

Detection and characterization of Tn2501, a transposon included within the lactose transposon Tn951.

The DNA sequence spanning coordinates 9.9 to 16.4 kilobases of the lactose transposon Tn951 ( Cornelis et al., Mol. Gen. Genet. 160:215-224, 1978) constitutes a transposable element by itself. Unlike Tn951 ( Cornelis et al., Mol. Gen. Genet. 184:241-248, 1981), this element, called Tn2501 , transposes in the absence of any other transposon. Transposition of Tn2501 proceeds through transient cointegration and duplicates 5 base pairs of host DNA. Tn2501 is flanked by nearly perfect inverted repeats (44 of 48), related to the inverted repeats of Tn21 ( Zheng et al., Nucleic Acids Res. 9:6265-6278, 1982). Unlike Tn21 , Tn2501 does not confer mercury resistance.

Base Sequence↗
PubMed1983

Study of the incompatibility and replication of the 70-kb virulence plasmid of Yersinia.

The 70-kb virulence plasmid, vir, from four Yersinia enterocolitica and one Y. pseudotuberculosis strains are incompatible with IncFI plasmids F'Lac and R386 while they are compatible with plasmids representing nine other incompatibility groups. Hybridization experiments carried out on one of these virulence plasmids showed that it contains the F incompatibility determinant D, incD. This determinant was cloned onto pACYC184 and the recombinant clone expressed incompatibility with F'Lac. We conclude that the incompatibility observed between F or R386 and the 70-kb virulence plasmid of Y. enterocolitica and Y. pseudotuberculosis is mediated by incD. Replication genes (rep) from the same plasmid were cloned independently in Escherichia coli. Rep and incD map on two different BamHI fragments. Surprisingly, the replicon isolated is not sensitive to inc D incompatibility. Apart from incD, vir and F share extremely little homology. In particular, there is no evidence for the presence of an F-like transfer operon on vir.

Calcium↗
PubMed1983

Haemoglobin measurement: the reliability of some simple techniques for use in a primary health care setting.

The reliability of five simple techniques for the estimation of blood haemoglobin levels was studied under laboratory conditions. The methods tested were the Sahli technique, the Tallqvist method, and the Lovibond comparator with undiluted blood, blood diluted in Drabkin solution, and blood diluted in modified Drabkin solution. The Lovibond comparator technique using undiluted blood was satisfactory in terms of precision and accuracy, and seems suitable for the monitoring of individual patients and for screening for anaemia. The other techniques were less reliable.A detailed procedure was worked out for estimation of the number of screening errors that can be expected with each of these techniques in any epidemiological situation.

Anemia↗
PubMed1983

[Genetic approach to the virulence of bacteria].

Bacteria only possess a single chromosome, but they also contain prophages, plasmids and transposons. The genetic basis for the pathogenic power of invasive and toxigenic bacteria only represents a small part of the total genotype of these bacteria. It is curious that although the plasmids and the prophages only represent about 1 p. cent of the total genome, they are disproportionately responsible for carrying the genetic information concerning virulence. The genetic study of virulence has revealed new mechanisms of pathogenic activity, for example the interaction of siderophores, adherence factors and haemolysins in pathogenic activity. Furthermore, the understanding of the genetic basis of virulence will lead to new methods of diagnosis and a new generation of vaccines.

Bacteria↗
PubMed1981

Transposon Tn951 (TnLac) is defective and related to Tn3.

Tn951 is flanked by two perfect inverted repeats of 41 bp which include the 38 bp sequence of the IR of Tn3. Tn951 also contains the last 100 bp of the tnpA gene but with at least two mutations. However, beyond nucleotide 137 the sequences diverge and hybridization experiments show that Tn951 lacks at least the first two thirds of the tnpA gene. In agreement with these observations Tn951 does not transpose by itself at a detectable frequency but can be complemented by the tnpA gene of Tn801 or Tn3. Tn501, Tn1721 and gamma delta do not complement Tn951 transposition. Transposition of Tn951 duplicates 5 bp of target DNA sequence.

Base Sequence↗
PubMed1981

Sequence relationships between plasmids carrying genes for lactose utilization.

Southern hybridization experiments carried out between pSC101::Tn951 DNA and lambda dlac DNA allowed the location and orientation of the lac operon within the transposon to be deduced. The same method was used to detect Tn951 on Lac plasmids from 11 independent isolates from three continents. None of these plasmids was found to carry an entire Tn951 sequence but they all contained lac genes homologous to the lac genes of Tn951. The lac operon of one of these plasmids was bordered by a sequence homologous to that found at the left-hand side of Tn951. It is concluded that the lac determinants of the Lac plasmids analysed and of Tn951 have evolved from a common ancestor but that the distribution of these determinants cannot be attributed to a spread of the transposon Tn951.

Bacteriophage lambda↗
PubMed1981

Plasmid-encoded lysine decarboxylation in Proteus morganii.

As a rule, Proteus morganii does not decarboxylate lysine. However, lysine-positive P. morganii strains have been recently described. We suspected a plasmid origin for this atypical character, and we analyzed 14 strains to study this question. Among these strains, 8 yielded lysine-negative segregants after acridine orange or ethidium bromide treatment, and 10 transferred their lysine-positive character to a recipient P. morganii strain. All of the 14 strains analyzed at least segregated or conjugated. Three lysine-positive transconjugants, in turn, segregated lysine-negative variants after ethidium bromide treatment. The eight wild-type lysine-positive strains that segregated lysine-negative subclones contained a large (35 to 45 megadaltons) plasmid detectable by agarose gel electrophoresis. Similarly, the 10 lysine-positive transconjugants contained a plasmid of the same size, whereas P. morganii 1000, the recipient strain, did not contain any detectable plasmid. The large plasmid clearly disappeared in 9 of 11 lysine-negative segregants analyzed. It is concluded that the lysine-positive character of these P. morganii strains is plasmid encoded, and the taxonomical implications are discussed.

Carboxy-Lyases↗
PubMed1980

Deletions and an inversion induced by a resident IS1 of the lactose transposon Tn951.

DNA-DNA filter binding tests, "Southern" blotting experiments and DNA heteroduplex analysis clearly show that Tn951 contains an IS1 element. This IS1-951 sequence is peculiar in that it does not contain the PstI cleavage site which is usually observed on E. coli derived IS1 elements. Nonetheless, IS1-951 induces deletions. This process is temperature dependent. One instance of an IS1-951 induced inversion was observed, the structure of which is compatible with the current models of transposition of IS elements.

Base Sequence↗
PubMed1980

Transposition of Tn951 (Tnlac) and cointegrate formation are thermosensitive processes.

The frequency of transposition of Tnlac to pGC200, an IncFII R plasmid, increased during the storage of the host strain. This result is explained by the fact that the transpositional event is temperature-dependent: it occurred readily when the host strain was grown at 30 degrees C but it was nearly undetectable when the host strain was grown and kept at 37 degrees C. Fusions between two different plasmids carrying Tnlac with pGC200 were also thermosensitive, suggesting a relation between cointegrate formation and transposition. Lactose did not influence the frequency of transposition of Tnlac.

DNA↗
PubMed1980

Expression of the lactose transposon Tn951 in Escherichia coli, Proteus and Pseudomonas.

The control of beta-galactosidase specified by the lactose transposon Tn951 (inserted into RP1 to give pGC9114) has been studied in Escherichia coli K12, Proteus mirabilis, Pseudomonas aeruginosa and Pseudomonas putida; in the first two species comparison could be made with Flac. In E. coli K12, the Tn951 and chromosomally encoded enzymes showed marked qualitative differences in regulatio, the former giving a substantially lower maximum induced level and induction ratio. Several parameters were slightly affected by strain background. In P. mirabilis, beta-galactosidase control determined by both Flac (in accord with earlier work) and pGC9114 was markedly different from E. coli in that maximal induced levels were about an order of magnitude lower and the induction ratio was reduced to 3 to 5. In Ps. aeruginosa and Ps. putida, Tn951-specified lac expression was qualitatively similar to that in P. mirabilis. Possible reasons for anomalous expression in Proteus and Pseudomonas are discussed.

DNA Transposable Elements↗