Microbiology & Immunology - Research Publications

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    Identification and isolation of slow-cycling glioma stem cells
    Furst, L ; Atkins, RJ ; Dinevska, M ; Stylli, SS ; Corcoran, NM ; Hovens, CM ; Mantamadiotis, T ; Vitale, I ; Manic, G ; Galluzzi, L (ELSEVIER ACADEMIC PRESS INC, 2022)
    Cancer stem cells are defined as low-abundance, quiescent cells and are considered a major cellular source of tumor recurrence following therapy, which identifies these cells as important therapeutic targets for difficult-to-treat cancers, including high-grade gliomas. By contrast to the highly proliferative bulk tumor cells, glioma stem cells (GSC) are slow-cycling, and therefore less sensitive to DNA damaging cytotoxic drugs. GSC are also less reliant on aerobic glycolytic metabolism, leading to inadequate clearing of GSC by chemotherapy and radiotherapy. The definition of GSC is based on the expression of specific stem cell protein markers. This method of GSC isolation is successful in isolating cell populations that can reliably recapitulate the tumor. However, cell populations that lack stem marker expression may also be capable of tumor recapitulation. Therefore, robust, reproducible methods for isolating GSC are required to identify and isolate cells with stem cell characteristics. Here, we provide a comprehensive and reproducible protocol for the isolation of slow-cycling GSC. Using this method, GSC isolated retain key characteristics of the cells in situ, including expression of genes associated with cell quiescence and invasive potential, compared to non-quiescent cell populations. Thus, isolation of GSC gated on cell proliferation offers a reliable alternative method for in vitro GSC identification, that adequately mirrors the physiological properties of GSC seen in vivo.
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    Glycopeptide-Centric Approaches for the Characterization of Microbial Glycoproteomes.
    Scott, NE (Springer US, 2022)
    Protein glycosylation is increasingly recognized as a common class of modifications within microbial species that can shape protein functions and the proteome at large. Due to this, there is an increasing need for robust analytical methods, which allow for the identification and characterization of microbial glycopeptides from proteome samples in a high-throughput manner. Using affinity-based enrichment (either hydrophilicity or antibody-based approaches) glycopeptides can easily be separated from non-glycosylated peptides and analyzed using mass spectrometry. By utilizing multiple mass spectrometry fragmentation approaches and open searching-based bioinformatic techniques, novel glycopeptides can be identified and characterized without prior knowledge of the glycans used for glycosylation. Using these approaches, glycopeptides within samples can rapidly be identified as well as quantified to understand how glycosylation changes in response to stimuli or how changes in glycosylation systems impact the glycoproteome. This chapter outlines a set of robust protocols for the initial preparation, enrichment, and analysis of microbial glycopeptides for both qualitative and quantitative glycoproteomic studies. Using these approaches, glycosylation events can be easily identified by researchers without the need for extensive manual analysis of proteomic datasets.
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    Evaluation of Human Circulating T Follicular Helper Cells in Influenza- and SARS-CoV-2-Specific B Cell Immunity.
    Koutsakos, M ; Kedzierska, K ; Nguyen, THO (Springer US, 2022)
    Generation of effective immune protection against viral infection and vaccination depends greatly on a successful engagement and stimulation of adaptive immune B cells and a specialized CD4+ T cell subset called T follicular helper cells (TFH cells). Since TFH cells primarily reside in lymphoid tissues, they can be challenging to study in human settings. However, a counterpart of these cells, circulating TFH (cTFH) cells, can be detected in peripheral blood. Assessment of cTFH cells serves as an informative marker of humoral responses following viral infection and vaccination and can be predictive of antibody titers. Here, we describe a comprehensive flow cytometry detection method for dissecting cTFH subsets and activation, together with the assessment of antibody-secreting cells (ASCs), from a small volume of human whole blood. This approach allows the investigation of cellular events that underpin successful immune responses following influenza and SARS-CoV-2 infection/vaccination in humans and is applicable to other viral disease settings.
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    Dendritic Cells and Their Roles in Anti-Tumour Immunity
    Shan Pang, E ; Macri, C ; Patton, T ; Bafit, M ; O’Keeffe, M ; Rajer, M ; Segelov, E (IntechOpen, 2020)
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    Antimicrobial resistance in healthcare, agriculture and the environment: the biochemistry behind the headlines
    Venter, H ; Henningsen, ML ; Begg, SL ; Venter, H (PORTLAND PRESS LTD, 2017)
    The crisis of antimicrobial resistance (AMR) is one of the most serious issues facing us today. The scale of the problem is illustrated by the recent commitment of Heads of State at the UN to coordinate efforts to curb the spread of AMR infections. In this review, we explore the biochemistry behind the headlines of a few stories that were recently published in the public media. We focus on examples from three different issues related to AMR: (i) hospital-acquired infections, (ii) the spread of resistance through animals and/or the environment and (iii) the role of antimicrobial soaps and other products containing disinfectants in the dissemination of AMR. Although these stories stem from three very different settings, the underlying message in all of them is the same: there is a direct relationship between the use of antimicrobials and the development of resistance. In addition, one type of antimicrobial could select for cross-resistance to another type and/or for multidrug resistance. Therefore, we argue the case for increased stewardship to not only cover clinical use of antibiotics, but also the use of antimicrobials in agriculture and stewardship of our crucially important biocides such as chlorhexidine.
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    Epidemiology & Public Health
    VEITCH, MGK ; HOGG, G ; YUNG, AP ; MCDONALD, MM ; SPELMAN, DS ; STREET, AC ; JOHNSON, PJ (Yung, McDonald, Spelman, Street & Johnson, 2001)