TY - JOUR A1 - Hacker, Jörg A1 - Rdest, Ursula A1 - Wintermeyer, E. A1 - Ludwig, B. T1 - Legiolysin, a New Hemolysin from L. pneumophila N2 - Legionella pneumophila generares exotoxins, cytolysins, proteases oc hemolysins that darnage host cells llke erythrocytes or rissue cu lrure cells. The gene for a new L. pneumophila hemolysin withour a proteolytic activiry was idemified, cloned in E. coli and sequenced. The gene producr was analysed by SDS-Polyacrylamide-gel-electrophoresis. N2 - Legionella pneum.ophila bildet Exoroxine, Zytolysine, Proteasen oder Hämolysine, die Wirtszellen wie Erythrozyten oder Animalzellen schädigen. Das Gen für ein neues L. pneumophila Hämolysin ohne proteolytische Aktivität wurde identifiziert, in E. coli kloniert und sequenziert. Das Genprodukt wurde durch SDS-Gelelcktropborese analysiert. KW - Hämolysin KW - Legionella pneumophila Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-73070 ER - TY - JOUR A1 - Hacker, Jörg A1 - Ott, Manfred A1 - Wintermeyer, Eva A1 - Ludwig, Birgit A1 - Fischer, Gunter T1 - Analysis of virulence factors of Legionella pneumophila. N2 - Legionella pneumophila, the causative agent of Legionnaires' disease is a facultative intracellular bacterium, which in the course of human infection multiplies in lung macrophages predominantly manifesting as pneumonia. The natural habitat of Legionella is found in sweet water reservoirs and man-made water systems. Virulent L. pneumophila spontaneously convert to an avirulent status at a high frequency. Genetic approaches have led to the identification of various L. pneumophila genes. The mip (macrophage infectivity potentiator) determinant remains at present the sole established virulence factor. The Mip protein exhibits activity of a peptidyl prolyl cis trans isomerase (PPiase), an enzyme which is able to bind the immunosuppressant FK506 and is involved in protein folding. The recently cloned major outer membrane protein (MOMP) could play a role in the uptake of legionellae by macrophages. Cellular models are useful in studying the intracellular replication of legionellae in eukaryotic cells. Human celllines and protozoan models are appropriate for this purpose. By using U 937 macrophage-like cells and Acanthamoeba castellanii as hosts, we could discriminate virulent and avirulent L. pneumophila variants since only the virulent strain was capable of intracellular growth at 37 oc. By using these systems we further demonstrated that a hemolytic factor cloned and characterized in our laboratory, legiolysin (lly), had no influence on the intracellular growth of L. pneumophila. KW - Legionella pneumophila Y1 - 1993 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-70620 ER - TY - JOUR A1 - Hacker, Jörg A1 - Ott, Manfred A1 - Blum, Gabriele A1 - Marre, Reinhard A1 - Heesemann, Jürgen A1 - Tschäpe, Helmut A1 - Goebel, Werner T1 - Genetics of Escherichia coli uropathogenicity: Analysis of the O6:K15:H31 isolate 536 N2 - E. coli strain 536 (06: K15: H31) isolated from a case of acute pyelonephritis, expresses S-fimbrial adhesins, P-related fimbriae, common type I fimbriae, and hemolysins. The respective chromosomally encoded determinants were cloned by constructing a genomic library of this strain. Furthermore, the strain produces the iron uptake substance, enterocheline, damages HeLa cells, and behaves in a serum-resistant mode. Genetic analysis of spontaneously arising non-hemolytic variants revealed that some of the virulence genes were physically linked to large unstable DNA regions, termed "pathogenicity islands", which were mapped in the respective positions on the E. coli K-12linkage map. By comparing the wild type strain and mutants in in vitro and in vivo assays, virulence features have been evaluated. In addition, a regulatory cross talk between adhesin determinants was found for the wild-type isolate. This particular mode of virulence regulation is missing in the mutant strain. KW - Escherichia coli KW - Genetik Y1 - 1992 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-71578 ER - TY - JOUR A1 - Hacker, Jörg A1 - Ott, M. A1 - Schmidt, G. A1 - Hull, R. A1 - Goebel, W. T1 - Molecular cloning of the F8 fimbrial antigen from Escherichia coli N2 - The genetic determinant coding for the Pspecific F8 fimbriae was cloned from · the chromosome of the Escherichia coli wild-type strain 2980 (018: K5: H5: FlC, F8). The F8 determinant was further subcloned into the Pstl site of pBR322 and a restriction map was established. In a Southern hybridization experiment identity between the chromosomally encoded F8 determinant of 2980 and its cloned Counterpart was demonstrated. The cloned F8 fimbriäe and those of the wild type strain consist of a protein subunit of nearly 20 kDa. F8 fimbriated strains were agglutinated by an F8 polyclonal antiserum, caused mannose-resistant hemagglutination and attached to human uroepi thellal cells. The cloned F8 determinant was weil expressed in a variety of host strains. KW - Infektionsbiologie KW - Escherichia coli KW - antigen KW - F8 fimbriae KW - gene cloning Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-59391 ER - TY - JOUR A1 - Hacker, Jörg A1 - Ott, M. A1 - Ludwig, B. A1 - Rdest, U. T1 - Intracellular survival and expression of virulence determinants of Legionella pneumophila N2 - Legionella pneumophila, the causative agent of Legionnaires' disease is able to live and multiply within macrophages as weil 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 fibro:blastic 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 sturlies will be necessary to determine the exact role of the cloned proteins in the pathogenesis of Legionella. Zusammenfassung: Intrazelluläres Überleben N2 - Intrazelluläres Überleben und Expression der Virulenzdeterminanten vonLegionella pneumophila. L. pneumophila, der Erreger der Legionärskrankheit, hat die Fähigkeit, sowohl in Makrophagen als auch in Protozoen zu leben und sich dort zu vermehret;t. Legionellen inhibieren die Fusion von Phagosom und Lysosom und hemmen die Ansäuerung des Phagosoms. Mit Hilfe von zwei unterschiedlichen Zellkultur-Systemen konnte gezeigt werden, daß Legionella-Stämme ihre Virulenz nach Laborpassage verlieren. Um die Mechanismen zu studieren, die für das intrazelluläre Überleben von Legionellen verantwortlich sind, haben wir eine Genbank des Legionella pneumophila-Stammes Philadelphia I in Escherichia coli K-12angelegt. Mit Hilfe der Cosmid-Klonierungstechnik war es möglich, fünf putative Virulenzfaktoren zu klonieren. Zwei von diesen Faktoren haben hämolytische Eigenschaften und drei sind Membran-assoziierte Proteine mit Molekulargewichten von 19, 26 und 60 kilodalton. Eines der hämolytischen Proteine, das Legiolysin, lysiert spezifisch humane Erythrozyten. Das zweite Hämolysin zeigt zusätzlich proteolytische Eigenschaften und schädigt sowohl Vero- als auch CHO-Zellen. Weitere Studien sind notwendig, um die Rolle der klonierten Proteine in der Pathogenese von Legionella exakt zu bestimmen. KW - Infektionsbiologie Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-59681 ER - TY - JOUR A1 - Hacker, Jörg A1 - Ott, M. A1 - Hof, H. T1 - Effects of low, subinhibitory concentrations of antibiotics on expression of a virulence gene cluster of pathogenic E. coli by using a wild-type gene fusion N2 - No abstract available KW - Infektionsbiologie Y1 - 1993 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-59874 ER - TY - JOUR A1 - Hacker, Jörg A1 - Knapp, S. A1 - Goebel, W. T1 - Spontaneous deletions and flanking regions of the chromosomal inherited hemolysin determinant of an Escherichia coli 06 strain N2 - The hemolytic Escherichia coli strain 536 (06) propagates spontaneous hemolysin- negative mutants at relatively high rates (10-3 to 10-4 ). One type of mutant (type I) lacks both secreted (external) and periplasmic (internal) hemolysin activity (HlYex - IHlYin -) and in addition shows no mannose-resistant hemagglutination (Mrh -), whereas the other type (type II) is HlYex -IHIYin + and Mrh +. The genetic determinants for hemolysin production (hly) and for mannose-resistant hemagglutination (mrh) of this strain are located on the chromosome. Hybridization experiments with DNA probes specific for various parts of the hly determinant reveal that mutants of type I have lost the total hly determinant, whereas those of type 11 lack only part of the hlyB that is essential for transport of hemolysin across the outer membrane. Using a probe that contains the end sequence of the plasmid pHly152-encoded hly determinant (adjacent to hlyB), we determined that a related sequence flanks also the hlyB-distal end of the chromosomal hly determinant of E. coli 536. In addition several other similar or even identical sequences are found in the vicinity of the hlyC- and the hlyB-distal ends of both the chromosomal and the plasmid hly determinants. Y1 - 1983 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-40260 ER - TY - JOUR A1 - Hacker, Jörg A1 - Kestler, H. A1 - Hoschützky, H. A1 - Jann, K. A1 - Lottspeich, F. A1 - Korhonen, T. K. T1 - Cloning and characterization of the S fimbrial adhesin (SfaII) complex of an Escherichia coli O18:K1 meningitis isolate N2 - S fimbrial adbesins (Sfa), which are able to recognize sialic acid-containing receptors on eukaryotic cells, are produced by Escherichia coli strains causing urinary tract infections or newbom meningitis. We recently described tbe cloning and molecular cbaracterization of a determinant, termed sftJI, from the chromosome of an E. coli urinary tract infection strain. Herewe present data conceming a S fimbria-specific gene duster, designated sfall, of an E. coli newbom meningitis strain. Like tbe Sfal complex, Sfall consists of tbe major subunit protein SfaA (16 kDa) and the minor subunit proteins SfaG (17 kDa), SfaS (15 kDa), and SfaH (29 kDa). The genes encoding tbe subunit proteins of Sfall were identified and sequenced. Their protein sequences were calculated from the DNA sequences and compared with tbose of the Sfal complex subunits. Altbough the sequences ofthe two major SfaA subunits ditf'ered markedly, tbe sequences ofthe minor subunits sbowed only a few amino acid exchanges (SfaG, SfaH) or were completely identical (SfaS). The introduction of a site-specific mutation into the gene sfaSII and subsequent analysis of an SfaS-negative clone indicated that sfaSII codes for the sialic acid-specific adhesin of tbe meninigitis isolate. These data were confirmed by tbe isolation and characterization of tbe SfaSII protein and the determination of its N-terminal amino acid sequence. The identity between the sialic acid-specific adhesins of Sfal and Sfall revealed that difl'erences between the two Sfa complexes with respect to tbeir capacities to agglutinate erythrocytes must result from sequence alterations of subunit proteins other tban SfaS. KW - Infektionsbiologie Y1 - 1993 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-59853 ER - TY - JOUR A1 - Hacker, Jörg A1 - Hughes, C. A1 - Hof, H. A1 - Goebel, W. T1 - Cloned hemolysin genes from Escherichia coli that cause urinary tract infection determine different levels of toxicity in mice N2 - After intraperitoneal injection of mice with Escherichia coli strains isolated from patients with urinary tract infections, the mortality due to hemolytic (Hly+) and nonhemolytic (Hiy-) isolates was 77 and 40%, respectively. Deletion of the chromosomal hemolysin (h/y) determinant in an E. co/i 06:K15:H31 urinary tract infection strain led to a significant reduction in toxicity for mice, and its reintroduction on a recombinant plasmid partially restored the original toxicity. Although introduction of the cloned plasmid pHiy152-encoded hly determinant into the Hly- E. coli 06 mutant strain increased toxicity by only a marginal degree, transformation with the cloned chromosomal hly determinants from two E. coli strains of serotypes 018ac:K5:H- and 075:K95:H? resulted in markedly greater toxicity, even exceeding that of the original Hly+ E. coli 06 wild-type strain. KW - Infektionsbiologie Y1 - 1983 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-59330 ER - TY - JOUR A1 - Hacker, Jörg A1 - Hof, H. A1 - Hughes, C. A1 - Goebel, W. T1 - Salmonella typhimurium strains carrying hemolysin plasmids and cloned hemolysin. genes from Escherichia coli N2 - Like all other Salmonella typhimurium strains examined, the smooth variants SF1397 (L T2) and 1366 and also their semi-rough and rough derivatives are non-haemolytic. Nevertheless, two haemolysin (Hly) plasmids of E. coli belonging to the inc groups incFllI,lv (pSU316) and incIz (pHly152) were able to be introduced into these strains by conjugation and stably maintained. A considerable percentage of the Hly+ transconjugants obtained had lost parts of their O-side chains, a result of selection for the better recipient capability of « semi-rough» variants rather than the direct influence of the Hly+ plasmids themselves. In contrast to the incF1lI1V plasmid pSU316, which exhibited higher conjugation rates with rough recipients, the incIz plasmid pHly152 was accepted best by smooth strains. Transformation with cloned E. coli haemolysin (hly) determinant was inefficient ( <10-8) for smooth strains, but 102-103 times higher for rough recipients, and was increased by the use of Salmonella-modified DNA. The transform ants and transconjugants were relatively stable and showed the same haemolytic activity as the E. coli donor strains. The virulence of the Hly+ smooth, semi-rough and rough S. typhimurium strains was tested in two mouse models, and neither the mortality rate nor the ability to multiply within the mouse spleen was influenced by the hly determinants. KW - Salmonella typhimurium KW - Plasmid KW - Haemolysin KW - Escherichia coli KW - Virulence Y1 - 1985 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-40309 ER -