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Measuring Phagocytosis of Aspergillus fumigatus Conidia by Human Leukocytes using Flow Cytometry
Published on: December 7, 2019
Automated quantification of the phagocytosis of Aspergillus fumigatus conidia by a novel image analysis algorithm
Kaswara Kraibooj1, Hanno Schoeler2, Carl-Magnus Svensson3
1Applied Systems Biology, Leibniz Institute for Natural Product Research and Infection Biology - Hans Knöll Institute Jena, Germany ; Faculty of Biology and Pharmacy, Friedrich Schiller University Jena Jena, Germany.
Abstract:
Studying the pathobiology of the fungus Aspergillus fumigatus has gained a lot of attention in recent years. This is due to the fact that this fungus is a human pathogen that can cause severe diseases, like invasive pulmonary aspergillosis in immunocompromised patients. Because alveolar macrophages belong to the first line of defense against the fungus, here, we conduct an image-based study on the host-pathogen interaction between murine alveolar macrophages and A. fumigatus. This is achieved by an automated image analysis approach that uses a combination of thresholding, watershed segmentation and feature-based object classification. In contrast to previous approaches, our algorithm allows for the segmentation of individual macrophages in the images and this enables us to compute the distribution of phagocytosed and macrophage-adherent conidia over all macrophages. The novel automated image-based analysis provides access to all cell-cell interactions in the assay and thereby represents a framework that enables comprehensive computation of diverse characteristic parameters and comparative investigation for different strains. We here apply automated image analysis to confocal laser scanning microscopy images of the two wild-type strains ATCC 46645 and CEA10 of A. fumigatus and investigate the ability of macrophages to phagocytose the respective conidia. It is found that the CEA10 strain triggers a stronger response of the macrophages as revealed by a higher phagocytosis ratio and a larger portion of the macrophages being active in the phagocytosis process.
Insights
A new automated image analysis method quantifies Aspergillus fumigatus (a fungus causing lung infections) interactions with immune cells. The CEA10 strain of A. fumigatus induced a stronger macrophage response than the ATCC 46645 strain.
Area of Science:
- Medical Mycology
- Immunology
- Computational Biology
Background:
- Aspergillus fumigatus is an opportunistic fungal pathogen causing severe invasive pulmonary aspergillosis in immunocompromised individuals.
- Alveolar macrophages are crucial in the initial defense against inhaled fungal conidia.
Purpose of the Study:
- To develop and apply an automated image analysis approach for studying host-pathogen interactions between murine alveolar macrophages and Aspergillus fumigatus.
- To quantitatively compare the interaction of two A. fumigatus wild-type strains (ATCC 46645 and CEA10) with macrophages.
Main Methods:
- Utilized automated image analysis combining thresholding, watershed segmentation, and feature-based object classification on confocal laser scanning microscopy images.
- Enabled segmentation of individual macrophages to compute the distribution of phagocytosed and adherent fungal conidia.
- Applied the framework to analyze interactions with A. fumigatus strains ATCC 46645 and CEA10.
Main Results:
- The automated analysis allowed for detailed quantification of cell-cell interactions and characteristic parameters.
- Macrophages exhibited a higher phagocytosis ratio and a greater proportion of active cells when interacting with the CEA10 strain compared to ATCC 46645.
- Demonstrated strain-specific differences in the host-pathogen interaction dynamics.
Conclusions:
- The developed automated image-based analysis provides a comprehensive framework for investigating host-pathogen interactions in fungal infections.
- Highlights the differential immune response elicited by A. fumigatus strains, with CEA10 showing increased interaction with macrophages.
- Offers a valuable tool for comparative investigations of fungal pathogen virulence and host defense mechanisms.
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