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Biomedical subjects

M Ott

Publications and source records attributed to M Ott.

At least 109 records · Page 6Linked to original sources
PubMed1991

[Intestinal permeability in patients with acquired immunodeficiency syndrome (AIDS)].

Intestinal permeability has been assessed as a parameter of the small intestinal barrier function in 33 patients with AIDS (WR 6), in 25 healthy volunteers, 32 patients with Crohn's disease, 18 patients with ulcerative colitis and in 12 patients with untreated coeliac disease. Permeability was measured by means of the 5 hour urinary excretion of lactulose and rhamnose after administration of these sugars (1 g rhamnose, 10 g lactulose) which allowed to calculate the urinary lactulose/rhamnose--ratio (L/R-r) as an index of intestinal permeability. Compared to the controls (L/R-r = 0.014 +/- 0.010) patients with AIDS had significantly increased lactulose/rhamnose--ratios (0.189 +/- 0.164; p less than 0.01) indicating abnormal permeation both of lactulose and rhamnose. This alteration of intestinal permeability was more pronounced than the significant increase of the L/R-r in the patients with either Crohn's disease or coeliac disease. However, intestinal permeability was not altered in the patients with ulcerative colitis. Among the patients with AIDS, the most abnormal permeability ratios were observed in two subjects with intestinal cryptosporidiosis. This investigation demonstrates by means of the "double sugar ratio", that intestinal permeability in patients with AIDS is highly abnormal, the impairment being even more abnormal than in patients with other small intestinal disease.

Acquired Immunodeficiency Syndrome↗
PubMed1991

The T-cell-receptor repertoire in the synovial fluid of a patient with rheumatoid arthritis is polyclonal.

We have analyzed the T-cell-receptor repertoire expressed in the synovial fluid of a patient with rheumatoid arthritis by using an inverse polymerase chain reaction. Total RNA was isolated from Ficoll-purified mononuclear cells and converted into circularized double-stranded cDNA. Specific amplification of alpha- and beta-chain variable regions (V alpha and V beta) was achieved with inverted alpha- and beta-chain constant region (C alpha and C beta) primer pairs, and the amplification products were cloned into phage vectors. A total of 78 alpha and 76 beta clones were sequenced, and 67 and 72 productively rearranged alpha and beta genes were identified, respectively. Thirty-one V alpha, 33 alpha-chain joining region (J alpha), 29 V beta, and 12 beta-chain joining region (J beta) gene segments were found in the productively rearranged clones, indicating that the T-cell repertoire expressed in the synovial fluid of this RA patient is highly heterogenous and polyclonal. Comparison of peripheral blood and synovial fluid repertoires showed that the most abundant V beta sequences, V beta 2.1 and V beta 3.1, were enriched in the inflamed joint by a factor of 2 to 3. It is possible that T cells expressing these V beta gene segments, which recognize bacterial superantigens, play a role in the disease.

Amino Acid Sequence↗
PubMed1991

Analysis of the variability of S-fimbriae expression in an Escherichia coli pathogen.

The uropathogenic Escherichia coli wild-type strain 536 produces S-fimbriae, P-related fimbriae and type I fimbriae. Using immuno-colony dot and ELISA techniques, variants were detected showing an increased degree of S-fimbrial production. It was demonstrated by immunofluorescence microscopy that in normal (wild-type) and hyper-S-fimbriated E. coli populations non-fimbriated cells also exist, and that the percentage of S-fimbriated and non-fimbriated bacteria was roughly identical in either population. Hyper-S-fimbriated variants could be stably maintained. The transition from wild-type to hyper-S-fimbriation, which occurs spontaneously, is markedly higher than vice versa. Southern blot analysis of the S fimbrial adhesin (sfa) determinants of normal and hyper-fimbriated strains revealed no marked difference in the gene structure.

Adhesins, Escherichia coli↗
PubMed1991

Intracellular survival and expression of virulence determinants of Legionella pneumophila.

Legionella pneumophila, the causative agent of Legionnaires' disease is able to live and multiply within macrophages as well as within protozoan organisms. Legionella strains inhibit phagosome-lysosome fusion and phagosome acidification. By using two different cell culture systems, one derived from human macrophages and the other from human embryo lung fibroblastic cells, it is demonstrated that Legionella strains lose their virulence following cultivation in the laboratory. In order to study the mechanisms involved in intracellular survival of Legionella a genomic library of strain Legionella pneumophila Philadelphia I was established in Escherichia coli K-12. By cosmid cloning technique we were able to clone five putative virulence factors, two of which exhibit hemolytic activities and three of which represent membrane-associated proteins of 19, 26 and 60 kilodalton. One of the hemolytic proteins, termed legiolysin, represents a new toxin which specifically lyses human erythrocytes. The other hemolysin exhibits proteolytic properties in addition and is cytolytic for Vero and CHO cells. Further studies will be necessary to determine the exact role of the cloned proteins in the pathogenesis of Legionella.

Bacterial Toxins↗
PubMed1991

Phenotype versus genotype of the 19 kD peptido-glycan associated protein of Legionella (PpIA), among Legionellae and other gram-negative bacteria.

The protein PpIA (19 kD) cloned from a genomic library of Legionella pneumophila, Philadelphia 1, represents a peptido-glycan associated outer membrane protein in recombinant E. coli K-12 and L. pneumophila. It exhibits distinct sequence homology to lipoproteins of Haemophilus influenzae and E. coli. A ppIA specific DNA probe generated by PCR was used in Southern hybridizations of chromosomal DNA of Legionella strains and other Gram-negative pathogens. Under conditions of high stringency, hybridization could only be observed in L. pneumophila isolates, but all other Legionella strains tested displayed hybridization under lower stringency. No signals appeared after hybridization of chromosomal DNA from a variety of other bacteria. Using anti-PpIA monospecific polyclonal antibodies in Western blots, it was demonstrated that PpIA related proteins of nearly the same size are found in all L. pneumophila isolates and in a variety of, but not all, the Legionella species analysed here.

Bacterial Outer Membrane Proteins↗
PubMed1991

Virulence determinants of Escherichia coli O6 extraintestinal isolates analysed by Southern hybridizations and DNA long range mapping techniques.

A total of 16 Escherichia coli O6 strains isolated from cases of extraintestinal infections were analysed for the genetic presence and phenotypic expression of fimbrial adhesins (P, S/FIC, type 1), aerobactin and hemolysin. In addition restriction fragment length polymorphisms (RFLPs) of Xbal-cleaved genomic DNA of seven selected strains, separated by orthogonal field alternation gel electrophoresis (OFAGE) were determined and virulence-associated DNA probes were used for Southern hybridization studies of the Xbal-cleaved genomic DNAs. The virulence characteristics and hybridization patterns obtained differed between the various isolates. In three isolates hemolysin genes and P fimbrial determinants were located on the same Xbal fragments. Furthermore, multiple copies of FIC determinants (foc) could be detected in two strains. Our data show that the new technique of pulse field electrophoresis together with Southern hybridization represents a powerful tool for the genetic analysis of pathogenic bacteria.

Adhesins, Escherichia coli↗
PubMed1991

Temperature-dependent expression of flagella in Legionella.

Legionella pneumophila, the causative agent of Legionnaires' disease, was analysed by electron microscopy for production of surface structures. Crystalline surface (S-) layers and fimbriae were not detected, but monotrichous flagellation was seen. Polyclonal antibodies specific for the 47 kDa flagellin subunit of L. pneumophila Philadelphia I were used in Western blots to confirm the presence of flagella subunits in various L. pneumophila strains tested, but the antiserum also reacted with flagellin subunits of L. micdadei, L. hackelia [serogroup (SG) 1 and SG2] and L. longbeachae (SG2). Flagellation of Legionellae was shown to be temperature regulated. When the growth temperature of virulent and avirulent of strain L. pneumophila Philadelphia I was shifted from 30 degrees C to either 37 or 41 degrees C, a decrease in the percentage of flagellated bacteria within the population was observed.

Bacterial Outer Membrane Proteins↗
PubMed1991

Analysis of Legionella pneumophila serogroup 6 strains isolated from a hospital warm water supply over a three-year period by using genomic long-range mapping techniques and monoclonal antibodies.

Over a period of 3 years, Legionella pneumophila serogroup 6 strains were isolated from warm water outlets and dental units in the Dental Faculty and from the Surgery and Internal Medicine Clinics at the University of Dresden, Dresden, Germany. In the bacteriological unit of the above-mentioned facility, L. pneumophila serogroups 3 and 12 were grown from warm water specimens. The medical facilities are located in separate buildings connected with a ring pipe warm water system. All L. pneumophila serogroup 6 strains isolated from the warm water supply reacted with a serogroup-specific monoclonal antibody, but not with two other monoclonal antibodies which are subgroup specific, reacting with other serogroup 6 strains. The NotI genomic profiles obtained by pulsed-field gel electrophoresis of 25 serogroup 6 strains isolated from the Dental Faculty over a 3-year period, 1 isolate from the Internal Medicine Clinic, and 4 strains from the Surgery Clinic were identical. Furthermore, all these strains hybridized with a 300-kb NotI fragment when a legiolysin (lly)-specific DNA probe was used. The NotI pattern, however, differed from those of six serogroup 6 strains of other origins, one serogroup 12 strain from the bacteriological unit, and another six unrelated strains of serogroups other than serogroup 6. L. pneumophila serogroup 6 strains which can be divided into only two subgroups by the use of monoclonal antibodies are differentiated in at least six NotI cleavage types obtained by pulsed-field electrophoresis.

Antibodies, Monoclonal↗
PubMed1991

Virulence patterns and long-range genetic mapping of extraintestinal Escherichia coli K1, K5, and K100 isolates: use of pulsed-field gel electrophoresis.

A total of 127 extraintestinal Escherichia coli strains of the capsule serotypes K1, K5, and K100 from human and animal sources were analyzed for DNA sequences specific for the genes for various adhesins (P fimbriae [pap] and P-related sequences [prs], S fimbriae [sfa]/F1C fimbriae [foc], and type I fimbriae [fim]), aerobactin (aer), and hemolysin (hly). The expression of corresponding virulence factors was also tested. Twenty-four selected strains were analyzed by long-range DNA mapping to evaluate their genetic relationships. DNA sequences for the adhesins were often found in strains not expressing them, while strains with hemolysin and aerobactin genes usually did express them. Different isolates of the same serotype often expressed different virulence patterns. The use of virulence-associated gene probes for Southern hybridization with genomic DNA fragments separated by pulsed-field gel electrophoresis revealed that a highly heterogeneous restriction fragment length and hybridization pattern existed even within strains of the same serotype. Long-range DNA mapping is therefore useful for the evaluation of genetic relatedness among individual isolates and facilitates the performance of precise molecular epidemiology.

Adhesins, Escherichia coli↗
PubMed1991

Distribution, expression, and long-range mapping of legiolysin gene (lly)-specific DNA sequences in legionellae.

The legiolysin gene (lly) cloned from Legionella pneumophila Philadelphia 1 confers the phenotypes of hemolysis and browning of the culture medium. An internal lly-specific DNA probe was used in Southern hybridizations for the detection of lly-specific DNA in the genomes of legionellae and other gram-negative pathogenic bacteria. Under conditions of high stringency, the lly DNA probe specifically reacted with DNA fragments from L. pneumophila isolates; by reducing stringency, hybridization was also observed for all other Legionella strains tested. No hybridization occurred with DNAs isolated from bacteria of other genera. The lly gene was mapped by pulsed-field gel electrophoresis to the respective genomic NotI fragments of Legionella isolates. By using antilegiolysin monospecific polyclonal antibodies in Western blots (immunoblots), Lly proteins could be detected only in L. pneumophila isolates.

Animals↗
PubMed1991

Pulsed field electrophoresis of genomic restriction fragments for the detection of nosocomial Legionella pneumophila in hospital water supplies.

Ten Legionella pneumophila strains isolated from different sources were analyzed according to their restriction fragment patterns obtained by cleavage of genomic DNA with NotI and SfiI and separation by pulsed field electrophoresis. Three L. pneumophila isolates from a nosocomial outbreak in Lübeck (Germany) and three other L. pneumophila strains independently isolated from a water tap located in the care unit where the patients were hospitalized exhibited identical restriction fragment profiles. Therefore, we concluded that these environmental specimens were the source of the Legionnaires disease. Another two isolates from patients and two strains from the environment, all unrelated to the outbreak described, showed different cleavage patterns.

Adult↗
PubMed1990

Identical genes for trimethoprim-resistant dihydrofolate reductase from Staphylococcus aureus in Australia and central Europe.

The nucleotide sequence of a 1.25 kb BglII/EcoRI fragment from the 34 kb trimethoprim (Tp)-resistant plasmid pABU1 of Staphylococcus aureus 157/4696, isolated in Zürich, was determined. It contained the entire Tp-resistant dihydrofolate reductase gene, 197 bp of the thymidylate synthetase, 395 bp of a truncated gene and 111 bp of IS257R1. With the exception of one single base pair at position of 862 the sequence of the whole fragment was identical to nucleotides 1633 to 2885 of the Tp-resistant transposon Tn4003 in plasmid pSK1 from an Australian S. aureus isolate. This suggests the worldwide dissemination of Tn4003 in multiresistant Staphylococci.

Amino Acid Sequence↗
PubMed1990

Genome analysis of Legionella ssp. by orthogonal field alternation gel electrophoresis (OFAGE).

Various Legionella isolates from different sources and origins were analysed by orthogonal field alternation gel electrophoresis of NotI cleaved genomic DNA. The genome of L. pneumophila Philadelphia I, the original isolate of the epidemics in 1976, exhibits only five NotI fragments. Two virulent derivatives, derived from L. pneumophila Philadelphia I, which were obtained by prolonged passage on artificial culture media, did not differ from their isogenic virulent strain according the NotI fragment pattern. By summing the lengths of the NotI fragments, the genome size of L. pneumophila Philadelphia I was calculated as approximately 3.9 Mb. Environmental L. pneumophila strains exhibited different NotI patterns, as did Legionella strains not belonging to the species pneumophila. The usefulness of DNA long range mapping of Legionella ssp. with NotI for epidemiology and evaluation of their evolutionary relationships is discussed.

Biological Evolution↗
PubMed1990

Deletions of chromosomal regions coding for fimbriae and hemolysins occur in vitro and in vivo in various extraintestinal Escherichia coli isolates.

Fimbrial adhesins and hemolysins contribute to pathogenicity of extraintestinal Escherichia coli isolates causing urinary tract infections (UTI), sepsis and new born meningitis (NBM). Using gene cloning techniques and pulse field electrophoresis in combination with Southern hybridizations it was demonstrated that the genetic determinants coding for P and 'P-related' fimbrial adhesins and hemolysins are closely linked on the chromosomes of different pathogenic E. coli wild-type isolates. For two UTI strains, 536 (O6:K15) and J96 (O4:K6), a co-deletion of the linked gene clusters coding for hemolysin and fimbriae was observed. The deleted DNA regions which also comprise flanking DNA sequences were termed 'pathogenicity DNA islands'. Such 'pathogenicity DNA islands' were also detected in the genome of O18:K1 isolates of OMP type 6 but were absent on the chromosomes of O18:K1 strains of OMP type 9. A mutant strain, 536-22 was selected from rat kidneys after intraurethral infection of animals with the wild-type parental strain 536. This particular isolate also shows deletions of 'pathogenicity islands' leading to a non-pathogenic phenotype. It is therefore concluded that excisions of 'pathogenicity islands' from chromosomes of pathogenic E. coli strains are not restricted to the laboratory but also occur in vivo. The generation of deletions may represent a general mechanism of bacterial virulence modulation.

Adhesins, Escherichia coli↗
PubMed1990

Complete genetic organization and functional aspects of the Escherichia coli S fimbrial adhesion determinant: nucleotide sequence of the genes sfa B, C, D, E, F.

The S fimbrial adhesin (sfa) determinant of E. coli comprises nine genes situated on a stretch of 7.9 kilobases (kb) DNA. Here the nucleotide sequence of the genes sfa B and sfa C situated proximal to the main structural gene sfaA is described. Sfa-LacZ fusions show that the two genes are transcribed in opposite directions. The isolation of mutants in the proximal region of the sfa gene cluster, the construction of sfa-phoA gene fusions and subsequent transcomplementation studies indicated that the genes sfa B and sfa C play a role in regulation of the sfa determinant. In addition the nucleotide sequence of the genes sfa D, sfa E and sfa F situated between the genes sfa A and sfa G responsible for S subunit proteins, were determined. It is suggested that these genes are involved in transport and assembly of fimbrial subunits. Thus the entire genetic organization of the sfa determinant is presented and compared with the gene clusters coding for P fimbriae (pap), F1C fimbriae (foc) and type I fimbriae (fim). The evolutionary relationship of fimbrial adhesion determinants is discussed.

Adhesins, Escherichia coli↗
PubMed1990

Use of a wild-type gene fusion to determine the influence of environmental conditions on expression of the S fimbrial adhesin in an Escherichia coli pathogen.

S fimbrial adhesins (Sfa) enable pathogenic Escherichia coli strains to bind to sialic acid-containing eucaryotic receptor molecules. In order to determine the influence of culture conditions on the expression of the sfa determinant in a wild-type strain, we fused the gene lacZ, coding for the enzyme beta-galactosidase, to the sfaA gene, responsible for the major protein subunit of S fimbriae. By using a plasmid which carries an R6K origin, the sfaA-lac hybrid construct was site-specifically integrated into the chromosome of the uropathogenic E. coli strain 536WT. The expression of lacZ, which was under the control of the sfa wild-type promoters, was now equivalent to the sfa expression of strain 536WT. With the help of this particular wild-type construct, it was demonstrated that the sfa determinant is better expressed on solid media than in liquid broth. The growth rate had a strong influence on Sfa expression under aerobic but not under anaerobic conditions. Production of Sfa was further regulated by catabolite repression, osmolarity, and temperature.

Adhesins, Escherichia coli↗