Pathogenetic aspects of Legionnaires' disease: interaction of Legionella pneumophila with cellular host defences.
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Humans↗
Biomedical subjects
C Rechnitzer
Publications and source records attributed to C Rechnitzer.
Effect of Legionella pneumophila cytotoxic protease on human neutrophil and monocyte function.
The extracellular metalloprotease of Legionella pneumophila, also called tissue-destructive protease or major secretory protein, has been proposed as one of the virulence factors of this organism. Considering the decisive role played by the phagocytic cells in host defense against Legionella infection, we investigated the effect of this protease on the function of human neutrophils and monocytes. L. pneumophila protease inhibited the chemotactic response of neutrophils to F-Met-Leu-Phe and zymosan-activated serum in a concentration-dependent and heat-labile manner. A direct effect of the protease on the chemotactic activity of neutrophils was demonstrated by the continued inhibition of neutrophil chemotaxis when the protease was removed following pre-incubation of the cells. In contrast, the enzyme had no effect on monocyte chemotaxis. The protease inhibited, also in a concentration-dependent and heat-labile manner, the binding of F-Met-Leu-Phe to both cell types. Neutrophil and monocyte oxidative burst response, as measured by superoxide release and chemiluminescence response, was not significantly affected by the enzyme. A slight enhancement of PMA-stimulated superoxide release was induced by the protease in both cell types. Lastly, the protease inhibited the killing of Listeria monocytogenes by neutrophils or monocytes. Inhibition of Listeria killing was concentration-dependent, heat-labile, and did not require the presence of the enzyme in the bactericidal assay. The inhibitory activity of L. pneumophila protease on neutrophil chemotaxis and on the listericidal activity of human neutrophils and monocytes demonstrated in this study provides evidence for a role of this enzyme in the pathogenesis of Legionnaires' disease.
Bacterial Proteins↗
Demonstration of the intracellular production of tissue-destructive protease by Legionella pneumophila multiplying within guinea-pig and human alveolar macrophages.
The major extracellular enzyme of Legionella pneumophila, a metalloprotease, has been proposed as a pathogenic factor in Legionnaires' disease due to its cytotoxic, tissue-destructive, and phagocyte-inhibitory properties. The relevance of these activities depends on the production of the protease during infection, i.e. by L. pneumophila multiplying intracellularly. In this study, L. pneumophila was demonstrated to produce protease in guinea-pig and human alveolar macrophages infected in vitro. After 24 h infection, approximately 0.1 to 0.2 micrograms of protease per 10(6) bacteria was measured by ELISA in culture supernatants and lysates of the infected cells, whereas no protease could be detected immediately after infection. Immunogold labelling using anti-protease antibody showed the enzyme to be located within phagosomes and distributed throughout the macrophages. Recent observations have shown that this protease could modify host defence mechanisms through inhibition of bacterial killing by neutrophils and monocytes. The intracellular production of the enzyme in infected macrophages demonstrated here further supports a role for the protease in the pathogenesis of Legionnaires' disease.
Animals↗
Prevalence of IgM antibodies to nine Legionella species in Icelandic children.
In order to evaluate the prevalence of antibodies to Legionella species among children in Iceland, a prospective study was conducted in 424 children aged 1 month to 12 years, 28 of whom had an acute respiratory tract infection. Antibody titers to L. pneumophila serogroup 1-6, L. bozemanii, L. dumoffii, and L. micdadei were measured by microagglutination technique. Seroreactivity to Legionella spp. was found in 30% of the children greater than 3 years of age and in 22% of all children. The majority of the children with legionella antibodies had no history of previous or present respiratory tract infection. The children with previous pneumonia or bronchial asthma did not show a higher seroreactivity to legionella than children without such a history. Our observations suggests that Icelandic children are frequently exposed to Legionella species or closely related bacteria.
Age Factors↗
Inhibition of human natural killer cell activity by Legionella pneumophila protease.
The present study was undertaken to define the effect of Legionella pneumophila protease on natural killer (NK) cell function in vitro. Lysis of target cells by human NK cells was determined using a 51Cr-release assay. The protease inhibited the NK cell cytolytic activity in a concentration- and time-dependent manner. The inhibitory effect of the protease was not affected by alpha interferon or interleukin 2. Legionella pneumophila protease partly inhibited the binding of effector cells to target cells as studied in a single cell agarose assay of monocyte-depleted cell populations. This effect of the protease on the binding of NK cells to target cells could interfere with the previously described enhanced NK cell activity induced by Legionella pneumophila. We could demonstrate in vitro inhibition of NK cell activity by Legionella pneumophila protease at very low concentrations, suggesting a possible relevance of this mechanism in the pathogenesis of Legionnaires' disease.
Cytotoxicity, Immunologic↗
Interleukin 6 primes human neutrophil and monocyte oxidative burst response.
Interleukin 6 (IL-6), a 26-kDa inducible protein, is a cytokine with multiple biological activities. This paper reports on the regulatory role of rIL-6 on the function of human polymorphonuclear and mononuclear leukocytes, a property not described previously. rIL-6 by itself did not exhibit any chemotactic activity and it could not activate these cells for an oxidative burst response. Preincubation of both cell types with rIL-6 at concentrations of 5 and 50 ng/ml primed the cells for enhanced generation of oxygen radicals following stimulation with the chemotactic peptide f-Met-Leu-Phe or the phorbol ester PMA. The enhancement of the oxidative burst response occurred both at the level of superoxide anion generation, an early step in the activation pathway, and at the level of the hydrogen peroxide-myeloperoxidase mediated response, a later step in the oxidative burst pathway. The priming ability was abolished by heat treatment of rIL-6 at 100 degrees C but not at 70 degrees C. Stimulation of B cell growth and immunoglobulin production combined with enhancement of oxidative burst response of phagocytic cells by IL-6 provide an effective mechanism of fighting against invading micro-organisms.
Chemotaxis↗
A rapid method for purification of homogeneous Legionella pneumophila cytotoxic protease using fast protein liquid chromatography.
A purification method was developed to isolate Legionella pneumophila cytotoxic protease in a form suitable for biological assays. Culture supernatant of a clinical isolate of L. pneumophila, Knoxville 1 strain, was used as the starting material. The protease was purified by FPLC on a Mono Q column followed by ultrafiltration. The isolated proteolytic enzyme has a specific activity of 90 azocasein units/mg protein and is a 42 kDa monomeric protein as determined by SDS-PAGE and gel filtration chromatography. It is heat-labile and toxic to a variety of cells e.g. McCoy, SIRC, HeLa, and rhabdomyosarcoma cells, baby hamster and green monkey kidney cells, and human embryonic lung fibroblasts.
Cell Line↗
Engulfment of the Philadelphia strain of Legionella pneumophila within pseudopod coils in human phagocytes. Comparison with other Legionella strains and species.
In this paper we report our ultrastructural studies of the early phagocytosis of two different strains of L. pneumophila serogroup (SG) 1 (Philadelphia 1 and Knoxville 1) and of L. micdadei. These bacteria replicate, in vivo as well as in vitro, in eukaryotic cells e.g. in monocytes and macrophages. Whether or not the mode of entry of these organisms in phagocytes contributes to their intracellular survival is presently unknown. Whilst internalization of bacteria of the Philadelphia 1 strain occurred within a pseudopod coil, organisms of the Knoxville 1 strain and L. micdadei were phagocytized in the classical manner, i.e. between pseudopods. No ultrastructural differences were observed between the two strains of L. pneumophila SG 1 whereas L. micdadei appeared as shorter rods with an extracellular layer of relatively low electron density. The phenomenon of coiling phagocytosis was not affected by heat-killing the bacteria or preopsonization with specific antibody. Formation of phagolysosomes was seen when cells of the Knoxville strain and L. micdadei were used but not with the Philadelphia strain. In our experiments, the occurrence of coiling phagocytosis was specific for the Philadelphia 1 strain of L. pneumophila and independent of bacterial virulence. Thus, it seems most unlikely that the coiling phenomenon plays any important role in the resistance of Legionella to the killing abilities of phagocytic cells.
Humans↗
The E. coli immunosorbent as used in serodiagnosis of Legionella infections studied by crossed immunoelectrophoresis.
In this study we investigated an immunosorbent, E. coli blocking fluid (BF), proposed for use in the Legionella Indirect Fluorescent Antibody Test (IFA). With crossed immunoelectrophoresis (CIE) of clinically relevant Legionella species, only one heat-stable antigen (no. 1) cross-reacted with the BF preparation. Patients' sera with elevated Legionella IFA titres did not react with this antigen in CIE. Out of 23 IFA positive patients' sera, six had titres lowered significantly to negative, when BF was applied as serum diluent for the titration (IFA BF negative sera). All six sera were negative in the micro agglutination test (MA). None of the IFA BF negative sera contained any Legionella precipitins in CIE, whereas nine out of the remaining 17 IFA BF positive sera unchanged by BF contained one or more precipitins. CIE results could not explain the effect of BF in Legionella IFA, and further studies are needed to sufficiently define the use of immunosorbents in diagnostic Legionella serology.
Antibodies, Bacterial↗
Defective monocyte oxidative metabolism in a child with non-pneumonic legionellosis.
A case of non-pneumonic legionellosis with gastrointestinal manifestations is described in a 13-year-old boy without premorbidity. A selective defect in monocyte oxidative metabolism was found and consistently demonstrated throughout the following year. We suggest that the observed monocyte deficiency may have facilitated the development of legionnaires disease and that otherwise normal monocyte function may have contributed to the favourable course of disease.
Adolescent↗
Modulation of human neutrophil and monocyte oxidative burst by Legionella pneumophila sonic extract.
The effect of Legionella pneumophila sonic extract on human neutrophil and monocyte oxidative burst was studied by superoxide anion release and luminol-enhanced chemiluminescence assays. Legionella pneumophila sonic extract by itself did not stimulate neutrophils and monocytes. The sonic extract at 8-2000 micrograms/ml primed neutrophils for enhanced superoxide release and, at 8-62.5 micrograms/ml, for enhanced chemiluminescence. Monocytes were only primed for enhanced chemiluminescence at very low extract concentrations (below 16 micrograms/ml). Monocyte superoxide release was suppressed by extract concentrations higher than 2000 micrograms/ml and the chemiluminescence response of neutrophils and monocytes by concentrations higher than 250 and 125 micrograms/ml, respectively. The priming activity was heat stable and present in fractions below 5 kDa. On the basis of these findings it is suggested that enhanced production of oxygen metabolites by neutrophils in contact with legionella components at low concentrations could contribute to the lung tissue damage seen in Legionnaires' disease, whereas the suppression of phagocyte oxidative burst by higher extract concentrations may be one of the mechanisms by which Legionella pneumophila survives intracellularly.
Endotoxins↗
Borrelia meningitis in Denmark.
We studied 23 patients with borrelia meningitis, a tick-borne spirochetosis increasingly recognized in Denmark. The disease showed a seasonal incidence from June to December. Twenty one patients experienced a painful sensory radiculitis, in 17 cases followed by a predominantly cranial mononeuritis multiplex mainly affecting the facial nerve (Bannwarth's syndrome). Three patients had focal CNS involvement in the form of a transverse myelitis, a focal encephalitis and a disseminated leucoencephalopathy, respectively. The CSF showed invariably lymphocytic pleocytosis and elevated spinal protein concentration. Measuring borrelia antibodies in the CSF seemed of higher diagnostic sensitivity than in serum. Penicillin therapy was effective even in cases with focal CNS lesions.
Adolescent↗
Priming of neutrophils for enhanced oxidative burst by sputum from cystic fibrosis patients with Pseudomonas aeruginosa infection.
Neutrophils accumulate in the lungs of cystic fibrosis (CF) patients and inflict tissue damage by release of oxygen radicals and proteases. Here we report on the ability of sputum to prime neutrophils for enhanced release of oxygen radicals. Sol phase was prepared by ultracentrifugation of sputum obtained from CF patients attending the CF Clinic, Rigshospitalet, Copenhagen. The oxidative burst response of neutrophils from healthy individuals was measured by oxygen consumption, superoxide production and chemiluminescence. Neutrophils were preincubated with sputum or buffer and then stimulated with f-Met-Leu-Phe or zymosan. Appropriate controls were included in the experiments. It was shown that neutrophils preincubated with CF sol phase and stimulated with f-Met-Leu-Phe generated a three- to five-fold higher chemiluminescence response than those preincubated with buffer. There was no enhancement of the response when zymosan was used for stimulation of the cells. Neutrophils incubated with sol phase alone exhibited no response. Attempts were made to identify and partially characterize the priming factor(s). It was found that the sputum samples contained bacterial endotoxins. The priming activity was resistant to heating at 100 degrees C for 15 min, and was present only in fractions with molecules larger than 100 KD. It is suggested that the priming factor(s) consist of bacterial endotoxins and/or immune complexes. Activation and enhanced release of oxygen radicals from neutrophils may play an important role in the host defence as well as pathogenesis of tissue damage in the lungs of these patients.
Cystic Fibrosis↗
[Legionella pneumophila pneumonia with lung abscess in a recipient of a cadaver kidney].
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Adult↗
Defective monocyte function in Legionnaires' disease complicating hairy cell leukaemia.
We describe a case of Legionnaires' disease in a 64-year-old man, in which hairy cell leukaemia was diagnosed after the onset of the infection. Immunological studies revealed a complete suppression of blood monocyte chemotactic and oxidative burst activities. We suggest that in hairy cell leukaemia both monocytopenia and defective functions of monocytes underlie the increased susceptibility to intracellular infections including Legionnaires' disease.
Diabetes Mellitus, Type 2↗
Effects of Legionella pneumophila sonicate on human neutrophil granulocyte and monocyte chemotaxis.
The interaction of L. pneumophila serogroup 1 sonic extract with human polymorphonuclear neutrophils and monocytes was studied in an in vitro chemotaxis assay. The sonicate showed heat-stable chemotactic activity towards neutrophils and monocytes. It also exhibited cytotoxicity for neutrophils but not for monocytes. Incubation and pretreatment with the sonicate at apparently non-toxic concentration resulted in inhibition of the chemotactic response of neutrophils to various chemoattractants while monocyte chemotaxis was unaffected. The inhibitory activity was reduced to one half by heat treatment of the sonicate. High endotoxic activity was demonstrated in both heated and non-heated extract. Only two antigens (nos 1 and 61) were identified in the sonicate exposed to 121 degrees C for 1 h. It is therefore suggested that the lipopolysaccharide (no. 61) of L. pneumophila could contribute to the chemotactic activity. The adverse effects exerted by L. pneumophila on neutrophils may be one of the mechanisms by which Legionella bacteria resist the normal phagocytic host defenses.
Antigens, Bacterial↗
Prevalence of Legionnaires' disease in pneumonia patients admitted to a Danish department of infectious diseases.
During a 14-month study period, 92 patients admitted to the University Clinic for Infectious Diseases with pneumonia were investigated to determine the prevalence and severity of Legionnaires' disease (LD). The diagnosis of LD was based on positive serology. Antibodies to 10 different legionella antigens--Legionella pneumophila serogroups 1-6, Fluoribacter (Legionella) bozemanae, F. dumoffii, F. gormanii, and Tatlockia (Legionella) micdadei--were measured by the microagglutination (MA) and indirect fluorescent antibody (IFA) techniques. LD was diagnosed in 22 patients showing a 4-fold or greater rise of MA titers. 10 patients showed a 4-fold or greater rise of IFA titer, 2 had standing high titer. One patient died. Legionella infection was the second most common cause of pneumonia. However, in half of the cases legionella infection occurred concomitantly with Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, Chlamydia psittaci, or viral infections. All 22 LD cases were sporadic. LD had been contracted abroad by 6 patients. Two of the legionella pneumonias were hospital-acquired. Half of the LD patients were older than 60 years. The majority of cases occurred during the winter months. Neither clinical chemistry parameters nor clinical features could distinguish LD from other types of pneumonia.
Adolescent↗
[Cerebrospinal borreliosis or Bannwarth's syndrome. A neurological infection transmitted by a wood tick].
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Adolescent↗