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IV004 Dormancy of Mycobacterium tuberculosis in vivo as a function of altered cell wall composition
Wissenschaftliches Programm 55. DGHM-Tagung 29. September-1. Oktober 2003 in Dresden Abstracts - Kurzvortr~ge
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Rheumatogenic group A streptococci aggregate collagen: a molecular basis for poststreptococcal rheumatic disease Dinkla, K. 1, Rohde, M.1; Chhatwal, G.S}; Talay, S.R.~ IGBF-German Research Centre for Biotechnology; Microbial Pathogenesis and Vaccine Research This study aimed to show a link between rheumatogenic group A streptococci and collagen binding. Acute rheumatic fever and heart disease are serious auto-immune sequelae of group A streptococcal infection. The mechanisms, however, by which certain group A streptococci but not others evoke auto-immune inflammatory processes, remains unresolved. In this study we show that serotype M3 and M18 streptococci isolated during outbreaks of rheumatic fever have the unique capability to bind and aggregate basement membrane collagen type IV. In radioactive pulldown experiments M3 protein was identified as collagenbinding factor of M3 streptococci, whereas collagenbinding of M18 isolates is mediated by hyaluronic acid capsule. Specific degradation of hyaluronic acid capsule revealed in complete loss of collagen-binding activity in radioactive binding experiments. In vivo mouse passage converts an unencapsulated and collagenbinding negative, serotype M1 strain into an encapsulated, collagen binding strain. We show that collagen binding enables S. pyogenes to colonize collagen matrix in vitro and in vivo. Coating of bacteria with collagen and fibronectin protects them from phagocytosis in vivo. Immunization with recombinant M3 protein induced an anti collagen type IV response in mice. Sera from patients suffering from acute rheumatic fever show significantly increased titers of anti collagen type IV antibodies in ELISA experiments compared to sera from healthy individuals.
Dormancy of Mycobacterium tuberculosis in vivo as a function of altered cell wall composition Ulrichs, T.1; Seller, p.2, Bandermann, S.2; Pradl, L.2, J6rg, S.2; Krenn, V?; Morawietz, L.3; Kaufmann, S.2; Aichele, p4 1Freie Universit~t; Medizinische Mikrobiologie, Institut fClr Infektionsmedizin 2MPI fClr Infektionsbiologie; Immunologie 3CharitY; Pathologie 4Universit~t Freiburg; Medizinische Mikrobiologie ZiehI-Neelsen (ZN) stain is the key technique to diagnose mycobacterial infections. However, a high percentage of patients exhibit positive signs of tuberculosis as indicated by pathology, culture of mycobacteria and PCR analysis and yet remain negative for ZN stain. Here, we present evidence that ZN negative specimens represent M. tuberculosis bacilli in a dormant state with distinct cell wall alterations: The classical cell wall composition-dependent ZN stain of M. tuberculosis in lung sections was gradually lost with ongoing persistence of infection both in mice and patients. In contrast, detection of mycobacteria by cell http://www.dghm.org
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Wissenschaftliches Programm 55. DGHM-Tagung 29. September-1. Oktober 2003 in Dresden Abstracts - Kurzvortr~ge wall composition-independent staining using a polyclonal anti-M, boris Bacille-Calmette-Gu6rin serum remained positive throughout persistence. These findings have important implications for diagnosis, and also for chemotherapy and development of vaccine strategies.
CD8+ T-lymphocytes are equipped to attract Mycobacterium tuberculosis-infected macrophages and kill the intracellular pathogen Stegelmann, F.1; Wagner, M.2; R011inghoff, M?; Stenger, S.4
1Klinische Mikrobiologie, Immunologie und Hygiene; Friedrich Alexander Universit~t Erlangen-NElrnberg 2Medizinische Klinik 3; Medizin 3Institut fElr Klinische Mikrobiologie, Immunologie und Hygiene; Medizin 4Erlangen; Klinische Mikrobiologie, Immunologie und Hygiene Granulysin is an antimicrobial peptide, which is stored in the granules of CDS+ T-lymphocytes (CTL). Similarly, the CC chemokine regulated upon activation normal T-cell expressed and secreted (RANTES) is stored in cytosolic granules of various lymphocyte subsets. To explore the possibility of a functional interaction between chemokines and antimicrobial peptides, we determined the expression and function of these molecules in freshly isolated human CTL. 70% (range 48-86%) of the cells expressed RANTES as determined by intracellular flow cytometry. In contrast, only 15% (range 5-25%) of CTL stored granulysin. Next we asked whether CTL express RANTES and granulysin selectively or simultaneously. We found, that the vast majority (>90%) of granutysinpositive cells also produced RANTES. This suggested, that RANTES and granulysin act in concert, whereby the function of RANTES could be to attract infected cells, which are then attacked by granulysin. Alternatively, RANTES could synergize with granulysin by exerting direct antimicrobial activity. To investigate these possibilities we measured the ability of recombinant RANTES to attract macrophages infected with a virulent strain of Mycobacterium tuberculosis. Using chemotaxis assays we found that RANTES preferentially attracts infected macrophages as compared to uninfected cells. In contrast RANTES (1100 ng/ml) did not limit the extracellular or intracellular growth of Mycobacterium tuberculosis in human alveolar macrophages. In summary, our findings indicate the existence of a subset of human CTL that stores molecules, which combine their biological functions to attract infected macrophages and subsequently kill the pathogen.
http://www.dghm.org
132
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Rheumatogenic group A streptococci aggregate collagen: a molecular basis for poststreptococcal rheumatic disease Dinkla, K. 1, Rohde, M.1; Chhatwal, G.S}; Talay, S.R.~ IGBF-German Research Centre for Biotechnology; Microbial Pathogenesis and Vaccine Research This study aimed to show a link between rheumatogenic group A streptococci and collagen binding. Acute rheumatic fever and heart disease are serious auto-immune sequelae of group A streptococcal infection. The mechanisms, however, by which certain group A streptococci but not others evoke auto-immune inflammatory processes, remains unresolved. In this study we show that serotype M3 and M18 streptococci isolated during outbreaks of rheumatic fever have the unique capability to bind and aggregate basement membrane collagen type IV. In radioactive pulldown experiments M3 protein was identified as collagenbinding factor of M3 streptococci, whereas collagenbinding of M18 isolates is mediated by hyaluronic acid capsule. Specific degradation of hyaluronic acid capsule revealed in complete loss of collagen-binding activity in radioactive binding experiments. In vivo mouse passage converts an unencapsulated and collagenbinding negative, serotype M1 strain into an encapsulated, collagen binding strain. We show that collagen binding enables S. pyogenes to colonize collagen matrix in vitro and in vivo. Coating of bacteria with collagen and fibronectin protects them from phagocytosis in vivo. Immunization with recombinant M3 protein induced an anti collagen type IV response in mice. Sera from patients suffering from acute rheumatic fever show significantly increased titers of anti collagen type IV antibodies in ELISA experiments compared to sera from healthy individuals.
Dormancy of Mycobacterium tuberculosis in vivo as a function of altered cell wall composition Ulrichs, T.1; Seller, p.2, Bandermann, S.2; Pradl, L.2, J6rg, S.2; Krenn, V?; Morawietz, L.3; Kaufmann, S.2; Aichele, p4 1Freie Universit~t; Medizinische Mikrobiologie, Institut fClr Infektionsmedizin 2MPI fClr Infektionsbiologie; Immunologie 3CharitY; Pathologie 4Universit~t Freiburg; Medizinische Mikrobiologie ZiehI-Neelsen (ZN) stain is the key technique to diagnose mycobacterial infections. However, a high percentage of patients exhibit positive signs of tuberculosis as indicated by pathology, culture of mycobacteria and PCR analysis and yet remain negative for ZN stain. Here, we present evidence that ZN negative specimens represent M. tuberculosis bacilli in a dormant state with distinct cell wall alterations: The classical cell wall composition-dependent ZN stain of M. tuberculosis in lung sections was gradually lost with ongoing persistence of infection both in mice and patients. In contrast, detection of mycobacteria by cell http://www.dghm.org
131
Wissenschaftliches Programm 55. DGHM-Tagung 29. September-1. Oktober 2003 in Dresden Abstracts - Kurzvortr~ge wall composition-independent staining using a polyclonal anti-M, boris Bacille-Calmette-Gu6rin serum remained positive throughout persistence. These findings have important implications for diagnosis, and also for chemotherapy and development of vaccine strategies.
CD8+ T-lymphocytes are equipped to attract Mycobacterium tuberculosis-infected macrophages and kill the intracellular pathogen Stegelmann, F.1; Wagner, M.2; R011inghoff, M?; Stenger, S.4
1Klinische Mikrobiologie, Immunologie und Hygiene; Friedrich Alexander Universit~t Erlangen-NElrnberg 2Medizinische Klinik 3; Medizin 3Institut fElr Klinische Mikrobiologie, Immunologie und Hygiene; Medizin 4Erlangen; Klinische Mikrobiologie, Immunologie und Hygiene Granulysin is an antimicrobial peptide, which is stored in the granules of CDS+ T-lymphocytes (CTL). Similarly, the CC chemokine regulated upon activation normal T-cell expressed and secreted (RANTES) is stored in cytosolic granules of various lymphocyte subsets. To explore the possibility of a functional interaction between chemokines and antimicrobial peptides, we determined the expression and function of these molecules in freshly isolated human CTL. 70% (range 48-86%) of the cells expressed RANTES as determined by intracellular flow cytometry. In contrast, only 15% (range 5-25%) of CTL stored granulysin. Next we asked whether CTL express RANTES and granulysin selectively or simultaneously. We found, that the vast majority (>90%) of granutysinpositive cells also produced RANTES. This suggested, that RANTES and granulysin act in concert, whereby the function of RANTES could be to attract infected cells, which are then attacked by granulysin. Alternatively, RANTES could synergize with granulysin by exerting direct antimicrobial activity. To investigate these possibilities we measured the ability of recombinant RANTES to attract macrophages infected with a virulent strain of Mycobacterium tuberculosis. Using chemotaxis assays we found that RANTES preferentially attracts infected macrophages as compared to uninfected cells. In contrast RANTES (1100 ng/ml) did not limit the extracellular or intracellular growth of Mycobacterium tuberculosis in human alveolar macrophages. In summary, our findings indicate the existence of a subset of human CTL that stores molecules, which combine their biological functions to attract infected macrophages and subsequently kill the pathogen.
http://www.dghm.org
132