164 research outputs found
The role of TcdB and TccC subunits in secretion of the photorhabdus Tcd toxin complex
No full textThe Toxin Complex (TC) is a large multi-subunit toxin encoded by a range of bacterial pathogens. The best-characterized examples are from the insect pathogens Photorhabdus, Xenorhabdus and Yersinia. They consist of three large protein subunits, designated A, B and C that assemble in a 5:1:1 stoichiometry. Oral toxicity to a range of insects means that some have the potential to be developed as pest control technology. The three subunit proteins do not encode any recognisable export sequences and as such little progress has been made in understanding their secretion. We have developed heterologous TC production and secretion models in E. coli and used them to ascribe functions to different domains of the crucial B+C sub-complex. We have determined that the B and C subunits use a secretion mechanism that is either encoded by the proteins themselves or employ an as yet undefined system common to laboratory strains of E. coli. We demonstrate that both the N-terminal domains of the B and C subunits are required for secretion of the whole complex. We propose a model whereby the N-terminus of the C-subunit toxin exports the B+C sub-complex across the inner membrane while that of the B-subunit allows passage across the outer membrane. We also demonstrate that even in the absence of the B-subunit, that the C-subunit can also facilitate secretion of the larger A-subunit. The recognition of this novel export system is likely to be of importance to future protein secretion studies. Finally, the identification of homologues of B and C subunits in diverse bacterial pathogens, including Burkholderia and Pseudomonas, suggests that these toxins are likely to be important in a range of different hosts, including man
Elucidating the in vivo targets of photorhabdus toxins in real-time using drosophila embryos
No full textThe outcome of any bacterial infection, whether it is clearance of the infecting pathogen, establishment of a persistent infection, or even death of the host, is as dependent on the host as on the pathogen (Finlay and Falkow 1989). To infect a susceptible host bacterial pathogens express virulence factors, which alter host cell physiology and allow the pathogen to establish a nutrient-rich niche for growth and avoid clearance by the host immune response. However survival within the host often results in tissue damage, which to some cases accounts for the disease-specific pathology. For many bacterial pathogens the principal determinants of virulence and elicitors of host tissue damage are soluble exotoxins, which allow bacteria to penetrate into deeper tissue or pass through a host epithelial or endothelial barrier. Therefore, exploring the complex interplay between host tissue and bacterial toxins can help us to understand infectious disease and define the contributions of the host immune system to bacterial virulence. In this chapter, we describe a new model, the Drosophila embryo, for addressing a fundamental issue in bacterial pathogenesis, the elucidation of the in vivo targets of bacterial toxins and the monitoring of the first moments of the infection process in real-time. To develop this model, we used the insect and emerging human pathogen Photorhabdus asymbiotica and more specifically we characterised the initial cross-talk between the secreted cytotoxin Mcf1 and the embryonic hemocytes. Mcf1 is a potent cytotoxin which has been detected in all Photorhabdus strains isolated so far, which can rapidly kill insects upon injection. Despite several in vitro tissue culture studies, the biology of Mcf1 in vivo is not well understood. Furthermore, despite the identification of many Photorhabdus toxins using recombinant expression in E. coli (Waterfield et al. 2008), very few studies address the molecular mechanism of action of these toxins in relation to specific immune responses in vivo in the insect model
Host–pathogen interactions : proline gives insect pathogens the green light
No full textHow does a quiescent symbiont of a nematode worm know when to turn nasty? Metabolic analysis and genetic knockouts confirm that model insect pathogens can sense L-proline in insect blood. This not only serves as a wake-up call, activating secondary metabolite virulence factors, but also provides an energy source for a metabolic shift appropriate for adaptation to the host environment
Drosophila embryos as model systems for monitoring bacterial infection in real time
No full textDrosophila embryos are well studied developmental microcosms that have been used extensively as models for early development and more recently wound repair. Here we extend this work by looking at embryos as model systems for following bacterial infection in real time. We examine the behaviour of injected pathogenic (Photorhabdus asymbiotica) and non-pathogenic (Escherichia coli) bacteria and their interaction with embryonic hemocytes using time-lapse confocal microscopy. We find that embryonic hemocytes both recognise and phagocytose injected wild type, non-pathogenic E. coli in a Dscam independent manner, proving that embryonic hemocytes are phagocytically competent. In contrast, injection of bacterial cells of the insect pathogen Photorhabdus leads to a rapid ‘freezing’ phenotype of the hemocytes associated with significant rearrangement of the actin cytoskeleton. This freezing phenotype can be phenocopied by either injection of the purified insecticidal toxin Makes Caterpillars Floppy 1 (Mcf1) or by recombinant E. coli expressing the mcf1 gene. Mcf1 mediated hemocyte freezing is shibire dependent, suggesting that endocytosis is required for Mcf1 toxicity and can be modulated by dominant negative or constitutively active Rac expression, suggesting early and unexpected effects of Mcf1 on the actin cytoskeleton. Together these data show how Drosophila embryos can be used to track bacterial infection in real time and how mutant analysis can be used to genetically dissect the effects of specific bacterial virulence factors
Photorhabdus asymbiotica as an insect and human pathogen
No full textPhotorhabdus asymbiotica is a species of bacterium that is pathogenic to humans whilst retaining the ability to infect insect hosts. Currently, there are two recognised subspecies, P. asymbiotica subsp. asymbiotica and P. asymbiotica subsp. australis with strains isolated from various locations in the USA, Australia, Thailand, Nepal and Europe. Like other species of Photorhabdus, P. asymbiotica subsp. australis was shown to form a symbiotic relationship with a Heterorhabditis nematode. In contrast to most strains of Photorhabdus luminescens, P. asymbiotica can grow at 37 °C and this is a defining factor in its ability to cause human disease. Insights into other adaptations it has undergone that have enabled host switching to occur have come from whole genome sequencing and transcriptomic studies. P. asymbiotica has a smaller genome compared to P. luminenscens with a lower diversity of insecticidal toxins. However, it has acquired plasmids and several pathogenicity islands in its genome. These encode genes with similarity to effectors or systems found in other known human pathogens such as Salmonella and Yersinia and are therefore likely to contribute to human pathogenicity. Of crucial importance to virulence is the fact that P. asymbiotica undergoes a large metabolic shift at the human host temperature
Diversity of Xenorhabdus and Photorhabdus spp. and their symbiotic entomopathogenic nematodes from Thailand
No full textXenorhabdus and Photorhabdus spp. are bacterial symbionts of entomopathogenic nematodes (EPNs). In this study, we isolated and characterized Xenorhabdus and Photorhabdus spp. from across Thailand together with their associated nematode symbionts, and characterized their phylogenetic diversity. EPNs were isolated from soil samples using a Galleria-baiting technique. Bacteria from EPNs were cultured and genotyped based on recA sequence. The nematodes were identified based on sequences of 28S rDNA and internal transcribed spacer regions. A total of 795 soil samples were collected from 159 sites in 13 provinces across Thailand. A total of 126 EPNs isolated from samples taken from 10 provinces were positive for Xenorhabdus (n = 69) or Photorhabdus spp. (n = 57). Phylogenetic analysis separated the 69 Xenorhabdus isolates into 4 groups. Groups 1, 2 and 3 consisting of 52, 13 and 1 isolates related to X. stockiae, and group 4 consisting of 3 isolates related to X. miraniensis. The EPN host for isolates related to X. stockiae was S. websteri, and for X. miraniensis was S. khoisanae. The Photorhabdus species were identified as P. luminescens (n = 56) and P. asymbiotica (n = 1). Phylogenenic analysis divided P. luminescens into five groups. Groups 1 and 2 consisted of 45 and 8 isolates defined as subspecies hainanensis and akhurstii, respectively. One isolate was related to hainanensis and akhurstii, two isolates were related to laumondii, and one isolate was the pathogenic species P. asymbiotica subsp. australis. H. indica was the major EPN host for Photorhabdus. This study reveals the genetic diversity of Xenorhabdus and Photorhabdus spp. and describes new associations between EPNs and their bacterial symbionts in Thailand
Colour, seeing, and seeing colour in medieval literature
No full textThis thesis re-approaches medieval literature in terms of its investment in visuality in general and chromatic perception in particular. The introduction raises the philosophical problem off-colour: its status as an object for science, role in perception, and relationship to language and meaning as expressed within inter-subjective evaluation. Two modes of discourse for colour studies of medieval literature are proposed: the phenomenological (from the philosophical tradition of such as Maurice Merleau-Ponty) seeking localised networks and patterns of inter-subjective, embodied, perceptual meanings and values; and linguistic (informed by the philosophical psychology and language philosophy of Ludwig Wittgenstein), focusing on the lexicalisation of colour experience and creation of semantic distinctions corresponding with changing colour concepts, which in turn shape individual perceptions (both first-hand experience and that of reading). Part One introduces key medieval ideas and theories pertaining to visual perception in general, and chromatic perception in particular. The authority for, and influence on medieval writers of Plato's Timaeus, Aristotle's De Anima and Parva Naturalia, and relevant biblical material is considered. Subsequent chapters explore Patristic and Neo-Platonist developments in extramissive thought, locating within this tradition the roots of a synthesis of natural philosophy with Christian theology that is found in later medieval thought and its dealings with perception and colour. A key movement in the theology of light in relation to colour is connected to the wider philosophical movement from largely "extramissive" to largely "intromissive" models of perception. This shift in theory and its significance for colour perception is explained in terms of the impact of Aristotle's material colour theory as found in De anima and the De sensti et sensato section of his Parva Naturalia from the late twelfth century onwards. The part concludes with a detailed study of the nineteenth chapter of Bartholomaeus Anglicus's thirteenth-century encyclopedia, De Proprietatibus Rerum, which provides access to an important range of ideas and sources relevant for accessing the medieval mind in its intellectualized perception of colour. Lastly, such philosophical and theological sources and ideas as are found in Part One are compared with relevant examples from literary texts, ranging from the Middle English poem, The Parliament of the Three Ages, to Christine de Pisan's Le Livre de la Cite des Dames. Part Two treats colour perception in relation to a particular medieval phenomenon: the rise of medieval heraldry and the armorial function of the herald. It considers the spiritual and secular ideologies of chivalry and their relationship to armorial displays as found portrayed and construed in various genres of chivalric literature. Texts under discussion include books of chivalry and arms from the early thirteenth to fourteenth centuries, such as those principally indebted to New Testament armorial allegory and motif (from writers such as Ramon Llull to Geoffrey de Charny), to later fourteenth-century treatises employing Aristotle’s De sensu et sensato to establish a secular hierarchy of chivalric colours. The study culminates with Part Three, offering responses to and discussions of particular medieval fictions in terms of their phenomenological, linguistic and intertextual treatment of colour perception. Medieval texts addressed include, amongst others, Le Roman de la Rose by Guillaume de Lorris and Jean de Meun, and four Middle English metrical romances: Sir Gowther, Sir Amadace, Sir Launfal and Sir Gawain and the Green Knight
The KdpD/KdpE two-component system : integrating K+ homeostasis and virulence
No full textThe two-component system (TCS) KdpD/KdpE, extensively studied for its regulatory role in potassium (K+) transport, has more recently been identified as an adaptive regulator involved in the virulence and intracellular survival of pathogenic bacteria, including Staphylococcus aureus, entero-haemorrhagic Escherichia coli, Salmonella typhimurium, Yersinia pestis, Francisella species, Photorhabdus asymbiotica, and mycobacteria. Key homeostasis requirements monitored by KdpD/KdpE and other TCSs such as PhoP/PhoQ are critical to survival in the stressful conditions encountered by pathogens during host interactions. It follows these TCSs may therefore acquire adaptive roles in response to selective pressures associated with adopting a pathogenic lifestyle. Given the central role of K+ in virulence, we propose that KdpD/KdpE, as a regulator of a high-affinity K+ pump, has evolved virulence-related regulatory functions. In support of this hypothesis, we review the role of KdpD/KdpE in bacterial infection and summarize evidence that (i) KdpD/KdpE production is correlated with enhanced virulence and survival, (ii) KdpE regulates a range of virulence loci through direct promoter binding, and (iii) KdpD/KdpE regulation responds to virulence-related conditions including phagocytosis, exposure to microbicides, quorum sensing signals, and host hormones. Furthermore, antimicrobial stress, osmotic stress, and oxidative stress are associated with KdpD/KdpE activity, and the system's accessory components (which allow TCS fine-tuning or crosstalk) provide links to stress response pathways. KdpD/KdpE therefore appears to be an important adaptive TCS employed during host infection, promoting bacterial virulence and survival through mechanisms both related to and distinct from its conserved role in K+ regulation
Writing and the rights of reality: usurpation and potentiality in Derrida, Plato, Nietzsche, and Beckett
No full textThe thesis critically evaluates Jacques Derrida's conferral of the rights of reality on writing, focussing on his theory of an arche-text in light of the speculative nature of this theory. The theory is initially considered in the context of Derrida's elucidation of the usurpatory status of writing within the Platonic and Nietzschean texts. This consideration reveals an admission of writing's usurpatory status by both writers while at the same time demonstrating their awareness of the intrinsically speculative nature of this view, the significance of writing lying in its ability to exteriorise the radically indeterminate status of consciousness m relation to reality rather than its ability to displace consciousness or reality The analyses, therefore, not only bring the Derridean hypothesis of a repressive or phonocentric metaphysical episteme into question but also exhibit the historical and philosophical role of potentiality in relation to writing, writing's ultimate significance lying in its capacity to exteriorise our existence as a mode of potentiality. Accordingly, in the second half of the thesis the Derridean theory of writing is countered with a specifically Aristotelian theory of the text as it is exhibited in the prose of Samuel Beckett, an author whose significance lies in his close alignment with Derridean theory within contemporary criticism. It is demonstrated that this identification has obviated an awareness of the significance of potentiality within the Beckettian text, his work consequently being appraised in the previously neglected context of Aristotelian metaphysics
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