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Day J.,Ohio State University | Schlesinger L.S.,Center for Microbial Interface Biology | Friedman A.,Ohio State University
Tuberculosis | Year: 2010

Due to the complexity of the immune response to a Mycobacterium tuberculosis infection, identifying new, effective therapies and vaccines to combat it has been a problematic issue. Although many advances have been made in understanding particular mechanisms involved, they have, to date, proved insufficient to provide real breakthroughs in this area of tuberculosis research. The term "Translational Systems Biology" has been formally proposed to describe the use of experimental findings combined with mathematical modeling and/or engineering principles to understand complex biological processes in an integrative fashion for the purpose of enhancing clinical practice. This opinion piece discusses the importance of using a Translational Systems Biology approach for tuberculosis research as a means by which to go forward with the potential for significant breakthroughs to occur. © 2009 Elsevier Ltd. All rights reserved.

Torrelles J.B.,Center for Microbial Interface Biology | Schlesinger L.S.,Center for Microbial Interface Biology
Tuberculosis | Year: 2010

Mycobacterium tuberculosis (the causal agent of TB) has co-evolved with humans for centuries. It infects via the airborne route and is a prototypic highly adapted intracellular pathogen of macrophages. Extensive sequencing of the M. tuberculosis genome along with recent molecular phylogenetic studies is enabling us to gain insight into the biologic diversity that exists among bacterial strains that impact the pathogenesis of latent infection and disease. The majority of the M. tuberculosis cell envelope is comprised of carbohydrates and lipids, and there is increasing evidence that these microbial determinants that are readily exposed to the host immune system play critical roles in disease pathogenesis. Studies from our laboratory and others have raised the possibility that M. tuberculosis is adapting to the human host by cloaking its cell envelope molecules with terminal mannosylated (i.e. Man-α-(1 → 2)-Man) oligosaccharides that resemble the glycoforms of mammalian mannoproteins. These mannosylated biomolecules engage the mannose receptor (MR) on macrophages during phagocytosis and dictate the intracellular fate of M. tuberculosis by regulating formation of the unique vesicular compartment in which the bacterium survives. The MR is highly expressed on alveolar macrophages (predominant C-type lectin on human cells) and functions as a scavenger receptor to maintain the healthiness of the lung by clearing foreign particles and at the same time regulating dangerous inflammatory responses. Thus M. tuberculosis exploits MR functions to gain entry into the macrophage and survive. Key biochemical pathways and mycobacterial determinants involved in the development and maintenance of the M. tuberculosis phagosome are being identified. The phylogenetic diversity observed in M. tuberculosis strains that impact its cell wall structure together with the genetic diversity observed in human populations, including those elements that affect macrophage function, may help to explain the extraordinary evolutionary adaptation of this pathogen to the human host. Major developments in these areas are the focus of this review. © 2010 Elsevier Ltd. All rights reserved.

Haghighat A.-C.,Center for Microbial Interface Biology | Seveau S.,Center for Microbial Interface Biology
Journal of Immunological Methods | Year: 2010

We describe an automated fluorescence microscopy-based assay that quantifies the invasion of mammalian cells by intracellular pathogens. Pathogens associated with host cell surfaces, intracellular pathogens and mammalian cells are directly counted based on their specific fluorescent labeling. Such approach utilizes automated image acquisition and processing, and is thus ideally suited for high-throughput analyses. This method was validated using Listeria monocytogenes as a model intracellular pathogen.

Azad A.K.,Center for Microbial Interface Biology | Knoell D.,Ohio State University | Schlesinger L.S.,Center for Microbial Interface Biology
Genes and Immunity | Year: 2013

Genetic variation in C-type lectins influences infectious disease susceptibility but remains poorly understood. We used allelic mRNA expression imbalance (AEI) technology for surfactant protein (SP)-A1, SP-A2, SP-D, dendritic cell-specific ICAM-3-grabbing non-integrin (DC-SIGN), macrophage mannose receptor (MRC1) and Dectin-1, expressed in human macrophages and/or lung tissues. Frequent AEI, an indicator of regulatory polymorphisms, was observed in SP-A2, SP-D and DC-SIGN. AEI was measured for SP-A2 in 38 lung tissues using four marker single-nucleotide polymorphisms (SNPs) and was confirmed by next-generation sequencing of one lung RNA sample. Genomic DNA at the SP-A2 DNA locus was sequenced by Ion Torrent technology in 16 samples. Correlation analysis of genotypes with AEI identified a haplotype block, and, specifically, the intronic SNP rs1650232 (30% minor allele frequency); the only variant consistently associated with an approximately twofold change in mRNA allelic expression. Previously shown to alter a NAGNAG splice acceptor site with likely effects on SP-A2 expression, rs1650232 generates an alternative splice variant with three additional bases at the start of exon 3. Validated as a regulatory variant, rs1650232 is in partial linkage disequilibrium with known SP-A2 marker SNPs previously associated with risk for respiratory diseases including tuberculosis. Applying functional DNA variants in clinical association studies, rather than marker SNPs, will advance our understanding of genetic susceptibility to infectious diseases. © 2013 Macmillan Publishers Limited.

Soni S.,Center for Microbial Interface Biology | Soni S.,Ohio State University | Ernst R.K.,University of Maryland, Baltimore | Muszynski A.,University of Georgia | And 5 more authors.
Frontiers in Microbiology | Year: 2010

Francisella tularensis is a CDC Category A biological agent and a potential bioterrorist threat. There is no licensed vaccine against tularemia in the United States. A long-standing issue with potential Francisella vaccines is strain phase variation to a gray form that lacks protective capability in animal models. Comparisons of the parental strain (LVS) and a gray variant (LVSG) have identified lipopolysaccharide (LPS) alterations as a primary change. The LPS of the F. tularensis variant strain gains reactivity to F. novicida anti-LPS antibodies, suggesting structural alterations to the O-antigen. However, biochemical and structural analysis of the F. tularensis LVSG and LVS LPS demonstrated that LVSG has less O-antigen but no major O-antigen structural alterations. Additionally, LVSG possesses structural differences in both the core and lipid A regions, the latter being decreased galactosamine modification. Recent work has identified two genes important in adding galactosamine (flmF2 and flmK) to the lipid A. Quantitative real-time PCR showed reduced transcripts of both of these genes in the gray variant when compared to LVS. Loss of flmF2 or flmK caused less frequent phase conversion but did not alter intramacrophage survival or colony morphology. The LVSG strain demonstrated an intramacrophage survival defect in human and rat but not mouse macrophages. Consistent with this result, the LVSG variant demonstrated little change in LD50 in the mouse model of infection. Furthermore, the LVSG strain lacks the protective capacity of F. tularensis LVS against virulent Type A challenge. These data suggest that the LPS of the F. tularensis LVSG phase variant is dramatically altered. Understanding the mechanism of blue to gray phase variation may lead to a way to inhibit this variation, thus making future F. tularensis vaccines more stable and efficacious. © 2010 Soni, Ernst, Muszynski, Mohapatra, Perry, Vinogradov, Carlson and Gunn.

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