Abstract (Expand)

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.

Authors: K. Kraibooj, Hanno Schoeler, C. M. Svensson, Axel Brakhage, Marc Thilo Figge

Date Published: 9th Jun 2015

Journal: Front Microbiol

Abstract (Expand)

Pathogens manipulate the cellular mechanisms of host organisms via pathogen-host interactions (PHIs) in order to take advantage of the capabilities of host cells, leading to infections. The crucial role of these interspecies molecular interactions in initiating and sustaining infections necessitates a thorough understanding of the corresponding mechanisms. Unlike the traditional approach of considering the host or pathogen separately, a systems-level approach, considering the PHI system as a whole is indispensable to elucidate the mechanisms of infection. Following the technological advances in the post-genomic era, PHI data have been produced in large-scale within the last decade. Systems biology-based methods for the inference and analysis of PHI regulatory, metabolic, and protein-protein networks to shed light on infection mechanisms are gaining increasing demand thanks to the availability of omics data. The knowledge derived from the PHIs may largely contribute to the identification of new and more efficient therapeutics to prevent or cure infections. There are recent efforts for the detailed documentation of these experimentally verified PHI data through Web-based databases. Despite these advances in data archiving, there are still large amounts of PHI data in the biomedical literature yet to be discovered, and novel text mining methods are in development to unearth such hidden data. Here, we review a collection of recent studies on computational systems biology of PHIs with a special focus on the methods for the inference and analysis of PHI networks, covering also the Web-based databases and text-mining efforts to unravel the data hidden in the literature.

Authors: S. Durmus, T. Cakir, A. Ozgur, Reinhard Guthke

Date Published: 9th Apr 2015

Journal: Front Microbiol

Abstract (Expand)

Time-lapse microscopy is an important technique to study the dynamics of various biological processes. The labor-intensive manual analysis of microscopy videos is increasingly replaced by automated segmentation and tracking methods. These methods are often limited to certain cell morphologies and/or cell stainings. In this paper, we present an automated segmentation and tracking framework that does not have these restrictions. In particular, our framework handles highly variable cell shapes and does not rely on any cell stainings. Our segmentation approach is based on a combination of spatial and temporal image variations to detect moving cells in microscopy videos. This method yields a sensitivity of 99% and a precision of 95% in object detection. The tracking of cells consists of different steps, starting from single-cell tracking based on a nearest-neighbor-approach, detection of cell-cell interactions and splitting of cell clusters, and finally combining tracklets using methods from graph theory. The segmentation and tracking framework was applied to synthetic as well as experimental datasets with varying cell densities implying different numbers of cell-cell interactions. We established a validation framework to measure the performance of our tracking technique. The cell tracking accuracy was found to be >99% for all datasets indicating a high accuracy for connecting the detected cells between different time points.

Authors: S. Brandes, Z. Mokhtari, F. Essig, Kerstin Hünniger, Oliver Kurzai, Marc Thilo Figge

Date Published: 8th Nov 2014

Journal: Med Image Anal

Abstract (Expand)

Candida albicans bloodstream infection is increasingly frequent and can result in disseminated candidiasis associated with high mortality rates. To analyze the innate immune response against C. albicans, fungal cells were added to human whole-blood samples. After inoculation, C. albicans started to filament and predominantly associate with neutrophils, whereas only a minority of fungal cells became attached to monocytes. While many parameters of host-pathogen interaction were accessible to direct experimental quantification in the whole-blood infection assay, others were not. To overcome these limitations, we generated a virtual infection model that allowed detailed and quantitative predictions on the dynamics of host-pathogen interaction. Experimental time-resolved data were simulated using a state-based modeling approach combined with the Monte Carlo method of simulated annealing to obtain quantitative predictions on a priori unknown transition rates and to identify the main axis of antifungal immunity. Results clearly demonstrated a predominant role of neutrophils, mediated by phagocytosis and intracellular killing as well as the release of antifungal effector molecules upon activation, resulting in extracellular fungicidal activity. Both mechanisms together account for almost [Formula: see text] of C. albicans killing, clearly proving that beside being present in larger numbers than other leukocytes, neutrophils functionally dominate the immune response against C. albicans in human blood. A fraction of C. albicans cells escaped phagocytosis and remained extracellular and viable for up to four hours. This immune escape was independent of filamentation and fungal activity and not linked to exhaustion or inactivation of innate immune cells. The occurrence of C. albicans cells being resistant against phagocytosis may account for the high proportion of dissemination in C. albicans bloodstream infection. Taken together, iterative experiment-model-experiment cycles allowed quantitative analyses of the interplay between host and pathogen in a complex environment like human blood.

Authors: Kerstin Hünniger, T. Lehnert, K. Bieber, R. Martin, Marc Thilo Figge, Oliver Kurzai

Date Published: 20th Feb 2014

Journal: PLoS Comput Biol

Abstract (Expand)

The ability to adapt to diverse micro-environmental challenges encountered within a host is of pivotal importance to the opportunistic fungal pathogen Candida albicans. We have quantified C. albicans and M. musculus gene expression dynamics during phagocytosis by dendritic cells in a genome-wide, time-resolved analysis using simultaneous RNA-seq. A robust network inference map was generated from this dataset using NetGenerator, predicting novel interactions between the host and the pathogen. We experimentally verified predicted interdependent sub-networks comprising Hap3 in C. albicans, and Ptx3 and Mta2 in M. musculus. Remarkably, binding of recombinant Ptx3 to the C. albicans cell wall was found to regulate the expression of fungal Hap3 target genes as predicted by the network inference model. Pre-incubation of C. albicans with recombinant Ptx3 significantly altered the expression of Mta2 target cytokines such as IL-2 and IL-4 in a Hap3-dependent manner, further suggesting a role for Mta2 in host-pathogen interplay as predicted in the network inference model. We propose an integrated model for the functionality of these sub-networks during fungal invasion of immune cells, according to which binding of Ptx3 to the C. albicans cell wall induces remodeling via fungal Hap3 target genes, thereby altering the immune response to the pathogen. We show the applicability of network inference to predict interactions between host-pathogen pairs, demonstrating the usefulness of this systems biology approach to decipher mechanisms of microbial pathogenesis.

Authors: L. Tierney, Jörg Linde, S. Muller, S. Brunke, J. C. Molina, Bernhard Hube, U. Schock, Reinhard Guthke, K. Kuchler

Date Published: 12th Mar 2012

Journal: Front Microbiol

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