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SURVIVAL OF MYCOBACTERIUM AVIUM SUBSPECIES PARATUBERCULOSIS IN THE POLYMORPHONUCLEAR LEUKOCYTES OF A CROHN'S DISEASE PATIENT

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Date Issued:
2004
Abstract/Description:
Mycobacterium avium subspecies paratuberculosis (map) is an intracellular pathogen that is known to parasitize macrophages and monocytes. Map infiltrates gastrointestinal tract host tissue where it is the known etiological agent of johne's disease in ruminants and implicated in the etiology of crohn's disease in humans. Map's ability to survive within macrophages enables it to disseminate throughout the rest of the host, possibly infecting other circulating blood leukocytes. In this study, the survival and fate of map strain atcc 43015 (human isolate) following phagocytosis was determined using in vitro murine macrophage cell line j774a.1 and polymorphonuclear cells (pmnc's) from five crohn's disease patients. Pmnc's from three healthy individuals and two ulcerative colitis patients, as well as escherichia coli (atcc 11303) and mycobacterium tuberculosis strain h37ra (atcc 25177), were included as controls (moi 10:1). Maturation of the phagosome was determined by evaluating the presence of stage specific markers on the surface of the phagosomal membrane. The endosomal protein, transferrin receptor, and the lysosomal marker, lamp-1, were then immunostained with cy-5 conjugated secondary antibodies, and colocalization of bacteria with each marker was evaluated separately using confocal scanning laser microscopy (cslm). In both tissue models, colocalization of viable map and m. Tuberculosis with the early endosomal marker, transferrin receptor occurred with an estimated five fold higher frequency than did association with the late lysosomal marker, lamp-1, as compared to live e. Coli, and all dead bacterial species. Using differential live/dead staining and fluorescent microscopy, survival of m. Tuberculosis and map was estimated at 85% and 79%, respectively compared to only 14% for e. Coli. The outcome was similar for both tissue culture and pmncs from all patients tested, suggesting that map and m. Tuberculosis can survive readily in both cell types, and regardless of the disease state of the host or the killing power of the cell. Map's survival appears to mimic m. Tuberculosis', suggesting the ability to resist phagolysosome fusion, by maintaining association with the early endosomes. Overall, the data confirms map virulence in host human blood leukocytes.
Title: SURVIVAL OF MYCOBACTERIUM AVIUM SUBSPECIES PARATUBERCULOSIS IN THE POLYMORPHONUCLEAR LEUKOCYTES OF A CROHN'S DISEASE PATIENT.
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Name(s): Rumsey, John, Author
Naser, Saleh, Committee Chair
University of Central Florida, Degree Grantor
Type of Resource: text
Date Issued: 2004
Publisher: University of Central Florida
Language(s): English
Abstract/Description: Mycobacterium avium subspecies paratuberculosis (map) is an intracellular pathogen that is known to parasitize macrophages and monocytes. Map infiltrates gastrointestinal tract host tissue where it is the known etiological agent of johne's disease in ruminants and implicated in the etiology of crohn's disease in humans. Map's ability to survive within macrophages enables it to disseminate throughout the rest of the host, possibly infecting other circulating blood leukocytes. In this study, the survival and fate of map strain atcc 43015 (human isolate) following phagocytosis was determined using in vitro murine macrophage cell line j774a.1 and polymorphonuclear cells (pmnc's) from five crohn's disease patients. Pmnc's from three healthy individuals and two ulcerative colitis patients, as well as escherichia coli (atcc 11303) and mycobacterium tuberculosis strain h37ra (atcc 25177), were included as controls (moi 10:1). Maturation of the phagosome was determined by evaluating the presence of stage specific markers on the surface of the phagosomal membrane. The endosomal protein, transferrin receptor, and the lysosomal marker, lamp-1, were then immunostained with cy-5 conjugated secondary antibodies, and colocalization of bacteria with each marker was evaluated separately using confocal scanning laser microscopy (cslm). In both tissue models, colocalization of viable map and m. Tuberculosis with the early endosomal marker, transferrin receptor occurred with an estimated five fold higher frequency than did association with the late lysosomal marker, lamp-1, as compared to live e. Coli, and all dead bacterial species. Using differential live/dead staining and fluorescent microscopy, survival of m. Tuberculosis and map was estimated at 85% and 79%, respectively compared to only 14% for e. Coli. The outcome was similar for both tissue culture and pmncs from all patients tested, suggesting that map and m. Tuberculosis can survive readily in both cell types, and regardless of the disease state of the host or the killing power of the cell. Map's survival appears to mimic m. Tuberculosis', suggesting the ability to resist phagolysosome fusion, by maintaining association with the early endosomes. Overall, the data confirms map virulence in host human blood leukocytes.
Identifier: CFE0000184 (IID), ucf:52838 (fedora)
Note(s): 2004-12-01
M.S.
Burnett College of Biomedical Sciences, Department of Molecular Biology and Microbiology
Masters
This record was generated from author submitted information.
Subject(s): Mycobacteria
polymorphonuclear leukocytes
Crohn's Disease
Persistent Link to This Record: http://purl.flvc.org/ucf/fd/CFE0000184
Restrictions on Access: campus 2005-01-31
Host Institution: UCF

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