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Journal articleAsai M, Li Y, Singh Khara J, et al., 2019,
Galleria mellonella: a novel infection model for screening potential anti-mycobacterial compounds against members of the Mycobacterium tuberculosis complex
, Frontiers in Microbiology, Vol: 10, ISSN: 1664-302XDrug screening models have a vital role in the development of novel antimycobacterial agents which are urgently needed to tackle drug-resistant tuberculosis (TB). We recently established the larvae of the insect Galleria mellonella (greater wax moth) as a novel infection model for the Mycobacterium tuberculosis complex. Here we demonstrate its use as a rapid and reproducible screen to evaluate antimycobacterial drug efficacy using larvae infected with bioluminescent Mycobacterium bovis BCG lux. Treatment improved larval survival outcome and, with the exception of pyrazinamide, was associated with a significant reduction in in vivo mycobacterial bioluminescence over a 96 hour period compared to the untreated controls. Isoniazid and rifampicin displayed the greatest in vivo efficacy and survival outcome. Thus G. mellonella, infected with bioluminescent mycobacteria, can rapidly determine in vivo drug efficacy, and has the potential to significantly reduce and/or replace the number of animals used in TB research.
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Book chapterHill P, Helaine S, 2019,
Antibiotic Persisters and Relapsing Salmonella enterica Infections.
, Persister Cells and Infectious Disease, Editors: Lewis, Publisher: Springer Nature, ISBN: 9783030252410Antibiotic persistence is defined as the ability of a subpopulation of bacteria within a clonal antibiotic-susceptible population to survive antibiotic treatment. Studies on antibiotic persistence have traditionally been carried out on bacteria cultured in laboratory media. However, over recent years, there has been a push to study antibiotic persisters in more physiologically relevant systems. Thus, the concept of antibiotic persistence during infection, which refers to the ability of a subpopulation of bacteria to survive combined host and antibiotic challenges, has emerged as a major new frontier of research. Here, we discuss the relevance and principles of this concept using relapsing Salmonella enterica infections as an example. We critically evaluate the clinical and experimental evidence for the existence and importance of antibiotic persisters in relapsing Salmonella infections; we outline our current understanding of the molecular mechanisms that enable successful antibiotic persistence during infection; and, finally, we discuss the challenges for this nascent field going forward.
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Journal articleMurphy P, Xu Y, Rouse SL, et al., 2019,
Functional 3D architecture in an intrinsically disordered E3 ligase domain facilitates ubiquitin transfer
<jats:title>Abstract</jats:title><jats:p>Post-translational modification of proteins with ubiquitin represents a widely used mechanism for cellular regulation. Ubiquitin is activated by an E1 enzyme, transferred to an E2 conjugating enzyme and covalently linked to substrates by one of an estimated 600 E3 ligases (1). RING E3 ligases play a pivotal role in selecting substrates and priming the ubiquitin loaded E2 (E2~Ub) for catalysis (2,3). RING E3 RNF4 is a SUMO targeted ubiquitin ligase (4) with important roles in arsenic therapy for cancer (4,5) and in DNA damage responses (6,7). RNF4 has a RING domain and a substrate recognition domain containing multiple SUMO Interaction Motifs (SIM<jats:sc>s</jats:sc>) embedded in a region thought to be intrinsically disordered (8). While molecular details of SUMO recognition by the SIMs (8–10) and RING engagement of ubiquitin loaded E2 (3,11–15) have been determined, the mechanism by which SUMO substrate is delivered to the RING to facilitate ubiquitin transfer is an important question to be answered. Here, we show that the intrinsically disordered substrate-recognition domain of RNF4 maintains the SIMs in a compact global architecture that facilitates SUMO binding, while a highly-basic region positions substrate for nucleophilic attack on RING-bound ubiquitin loaded E2. Contrary to our expectation that the substrate recognition domain of RNF4 was completely disordered, distance measurements using single molecule Fluorescence Resonance Energy Transfer (smFRET) and NMR paramagnetic relaxation enhancement (PRE) revealed that it adopts a defined conformation primed for SUMO interaction. Mutational and biochemical analysis indicated that electrostatic interactions involving the highly basic region linking the substrate recognition and RING domains juxtaposed those regions and mediated substrate ubiquitination. Our results offer insight into a key step in substrate ubiquitination by a membe
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Journal articleCerny O, Holden D, 2019,
Salmonella SPI-2 type III secretion system-dependent inhibition of antigen presentation and T cell function
, Immunology Letters, Vol: 215, Pages: 35-39, ISSN: 0165-2478Salmonella enterica serovars infect a broad range of mammalian hosts, including humans, causing both gastrointestinal and systemic diseases. Effective immune responses to Salmonella infections depend largely on CD4+ T cell activation by dendritic cells (DCs). Bacteria are internalised by intestinal DCs and respond by translocating effectors of the Salmonella pathogenicity island 2 (SPI-2) type III secretion system (T3SS) into host cells. In this review, we discuss processes that are hijacked by SPI-2 T3SS effectors and how this affects DC biology and the activation of T cell responses.
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Journal articleBacigalupe R, Angeles Tormo-Mas M, Penades JR, et al., 2019,
A multihost bacterial pathogen overcomes continuous population bottlenecks to adapt to new host species
, Science Advances, Vol: 5, ISSN: 2375-2548While many bacterial pathogens are restricted to single host species, some have the capacity to undergo host switches, leading to the emergence of new clones that are a threat to human and animal health. However, the bacterial traits that underpin a multihost ecology are not well understood. Following transmission to a new host, bacterial populations are influenced by powerful forces such as genetic drift that reduce the fixation rate of beneficial mutations, limiting the capacity for host adaptation. Here, we implement a novel experimental model of bacterial host switching to investigate the ability of the multihost pathogen Staphylococcus aureus to adapt to new species under continuous population bottlenecks. We demonstrate that beneficial mutations accumulated during infection can overcome genetic drift and sweep through the population, leading to host adaptation. Our findings highlight the remarkable capacity of some bacteria to adapt to distinct host niches in the face of powerful antagonistic population forces.
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Journal articleGonzalez D, Mavridou D, 2019,
Making the best of aggression, the many dimensions of bacterial toxin regulation
, Trends in Microbiology, Vol: 27, Pages: 897-905, ISSN: 0966-842XMost bacteria use toxins to exclude competitors.As the synthesis and delivery of these moleculesentail considerablecostsfor the producers, theirexpressionis tightly regulated, often by molecular systems detecting physiological stressesor environment-specific cues.However, the ecological connection between such systemsand competitive behaviorsis not always clear. Here, we review the regulation of antibacterial toxins and propose a conceptual framework organizing the decision-making processes controlling toxin production. As bacteria are unable to precisely identify their competitors,we argue that toxin regulation primarily responds to cues directly or indirectly associated with the presence of competing strains. The density and fitnessof the producing populational so play a role in the decision-making process.Overall, we contendthat optimal toxin production strategies involvemonitoring of both self and foe.
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Journal articleMullineaux-Sanders C, Sanchez-Garrido J, Hopkins EGD, et al., 2019,
<i>Citrobacter rodentium</i>-host-microbiota interactions: immunity, bioenergetics and metabolism
, NATURE REVIEWS MICROBIOLOGY, Vol: 17, Pages: 701-715, ISSN: 1740-1526- Author Web Link
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PatentMarchesi JR, McDonald JAK, Mullish BH, 2019,
Clostridioides difficile
The invention relates to Clostridioides difficile, and in particular to compounds, polypeptides and mixtures for the treatment of C. difficile infections. The invention also relates to nucleic acids, vectors comprising these nucleic acids and microorganisms for the treatment of C. difficile infections, and to methods of identifying and matching faecal microbiota transplant (FMT) donors to FMT recipients.
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Journal articleLim C, Ha KP, Clarke R, et al., 2019,
Identification of a potent small-molecule inhibitor of bacterial DNA repair that potentiates quinolone antibiotic activity in methicillin-resistant Staphylococcus aureus
, Bioorganic and Medicinal Chemistry, Vol: 27, Pages: 1-7, ISSN: 0968-0896The global emergence of antibiotic resistance is one of the most serious challenges facing modern medicine. There is an urgent need for validation of new drug targets and the development of small molecules with novel mechanisms of action. We therefore sought to inhibit bacterial DNA repair mediated by the AddAB/RecBCD protein complexes as a means to sensitize bacteria to DNA damage caused by the host immune system or quinolone antibiotics. A rational, hypothesis-driven compound optimization identified IMP-1700 as a cell-active, nanomolar potency compound. IMP-1700 sensitized multidrug-resistant Staphylococcus aureus to the fluoroquinolone antibiotic ciprofloxacin, where resistance results from a point mutation in the fluoroquinolone target, DNA gyrase. Cellular reporter assays indicated IMP-1700 inhibited the bacterial SOS-response to DNA damage, and compound-functionalized Sepharose successfully pulled-down the AddAB repair complex. This work provides validation of bacterial DNA repair as a novel therapeutic target and delivers IMP-1700 as a tool molecule and starting point for therapeutic development to address the pressing challenge of antibiotic resistance.
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Journal articleFurniss C, Dortet L, Bolland W, et al., 2019,
Detection of colistin resistance in Escherichia coli using the MALDI Biotyper Sirius mass spectrometry system
, Journal of Clinical Microbiology, Vol: 57, Pages: 1-7, ISSN: 0095-1137Polymyxin antibiotics are a last-line treatment for multidrug-resistant Gram-negative bacteria. However, the emergence of colistin resistance, including the spread of mobile mcr genes, necessitates the development of improved diagnostics for the detection of colistin-resistant organisms in hospital settings. The recently developed MALDIxin test enables detection of colistin resistance by MALDI-TOF mass spectrometry in less than 15 minutes, but is not optimized for the mass spectrometers commonly found in clinical microbiology laboratories. In this study, we adapted the MALDIxin test for the MALDI Biotyper Sirius MALDI-TOF mass spectrometry system (Bruker Daltonics). We optimized the sample preparation protocol using a set of 6 MCR-expressing Escherichia coli clones and validated the assay with a collection of 40 E. coli clinical isolates, including 19 confirmed MCR producers, 12 colistin-resistant isolates which tested negative for commonly encountered mcr genes (i.e. likely chromosomally-resistant isolates) and 9 polymyxin-susceptible isolates. We calculated Polymyxin resistance ratio (PRR) values from the acquired spectra; a PRR value of zero, indicating polymyxin susceptibility, was obtained for all colistin-susceptible E. coli isolates, whereas positive PRR values, indicating resistance to polymyxins, were obtained for all resistant strains independent of the genetic basis of resistance. Thus, we report a preliminary feasibility study showing that an optimized version of the MALDIxin test, adapted for the routine MALDI Biotyper Sirius, provides an unbiased, fast, reliable, cost-effective and high-throughput way of detecting colistin resistance in clinical E. coli isolates.
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