NIHRDataInsights/HRCSData huggingface dataset:
Title: Regulatory role of non-coding RNAs in herpesvirus infections
Funder: Medical Research Council

Abstract: All herpesviruses share the ability to persist for life in infected individuals thus posing the risk of reactivation and disease. The eight human herpesviruses inflict a substantial burden of disease and economic costs in immunosuppressed patients and in pregnancy. Our lack of understanding of the molecular mechanisms governing viral gene expression and manipulation of host gene expression significantly hinders our ability to deal with the associated diseases and has obstructed the development of new therapies. Here I propose two closely related projects which ask three major questions. 1) How do herpesviruses modulate cellular and viral gene expression? 2) What is the contribution and function of herpesvirus miRNAs? 3) Can viral miRNAs serve as targets for novel antiviral drugs? The low temporal resolution of standard gene expression profiling and the inability to precisely measure RNA decay has severely hampered our understanding of the regulation of gene expression during virus infections. We developed a novel system for nascent transcriptome analysis (NTA) to overcome all of these problems We now combine NTA with next-generation sequencing to study regulation of gene expression during lytic MCMV and HCMV infection. Similar approaches will be applied to the ?- and ?-herpesvirus family (HSV-1, MHV68). Thereby, we will provide a comprehensive picture of the regulation of gene expression in herpesvirus infection. SILAC proteomics available at the Babraham Institute will support NTA data at whole proteome level but will also elucidate the effects of viral miRNAs on target protein synthesis. In close collaboration with a number of international groups my group established the infection of the mouse with the murine cytomegalovirus (MCMV) to study the function of cytomegalovirus miRNAs in vivo. Technologies like traceless-mutagenesis of viral genomes, miRNA target identification by RIP-Chip as well as in vivo experiments are well established. We will now study how viral miRNAs concert with viral proteins to provide a favourable environment for these viruses in vivo and how this knowledge can be used against them. Viral miRNA research is a highly competitive field of research. Based on the advantage of a timely start and well established collaborations with leading groups in the field I am confident that both of these studies are straightforward. The aims are realistically achievable and the outcome will greatly improve our understanding of herpesvirus biology in general. Most importantly, the methodological approaches are easily transferred to any other cellular process opening up a plethora of opportunities for the future.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION
Title: LEAD SERIES DEVELOPMENT & OPTIMISATION OF A NEW DRUG AGAINST ACTIVE AND LATENT TUBERCULOSIS
Funder: Medical Research Council

Abstract: A major failure of current tuberculosis (TB) therapies is that they predominantly target replicating Mycobacterium tuberculosis (Mtb) but are unable to sterilize slow growing (dormant) Mtb, leading to protracted treatment regimes and the development of drug resistance. We propose to generate a new drug against TB that is able to mitigate the shortcomings of current therapies, leading to improved treatment outcomes. Our strategy is to target the Mtb respiratory chain, specifically NADH:menaquinone oxidoreductase (ndh). This target is essential for the survival of replicating, dormant and drug resistant Mtb, and it is absent in humans. Over the past 2 years we have successfully progressed from target validation/hit identification to the discovery of novel potent (nM) inhibitors of ndh with corresponding potent (nM) in vitro sterilization activity against replicating and dormant Mtb. This proposed schedule of work is to advance these early leads through to late lead development and optimization in readiness for candidate selection.


Research activity codes: 5.1 PHARMACEUTICALS
Health categories: INFECTION
Title: Defining hotspots of malaria transmission
Funder: Medical Research Council

Abstract: Malaria transmission is spatially heterogeneous, and groups of homesteads that form hotspots or clusters of transmission can be identified. The presence of these hotspots makes malaria control measures less effective than they might be. However, adding targeted interventions to interrupt these hotspots will be highly effective. At present, we lack detailed epidemiological descriptions of the properties of hotspots and the ways in which they might be identified by malaria control programmes. Furthermore, in order to rationally design targeted interventions, we need to understand their transmission dynamics. For example, we need to know how much transmission in the surrounding area results from the hotspot, and how focal the point source is. I will analyse 19 years of historical data on severe malaria, mild malaria and asymptomatic infection in Kilifi, Kenya. I will use datasets from cohorts under active surveillance in the field, and passive dispensary and hospital level surveillance, to describe the spatial and temporal limits of individual clusters of transmission, and the epidemiological markers of them. I will obtain external validation of my findings by collaborations with investigators collecting spatial data on malaria cases in Africa, including the Gambia and Indonesia. In collaboration with Dominic Kwiatkowski in the Wellcome Trust Sanger Institute, I will conduct detailed genotyping studies to assign a bar-code to parasites. High resolution spatial and genotyping data will be combined to accurately identify transmission in and around hotspots, in order to predict the likely outcomes of intervening in hotspots. I will use a descriptive statistical approach for my primary analysis, but will also collaborate with Gil McVean (Oxford University), Dave Smith (Florida University) and Azra Ghani (Imperial College) to conduct post hoc analyses of the population genetic structure, potential indirect effects of interventions and Bayesian approaches to cluster determination, respectively.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT;2.4 SURVEILLANCE AND DISTRIBUTION
Health categories: INFECTION
Title: The microbiome of the helminth infected host: Implications for immunity and intestinal homeostasis
Funder: Medical Research Council

Abstract: Worm parasites (helminthes) are capable of infecting the gastrointestinal tract a broad range of animals and are often associated with pronounced effects on the health of the host. Although interactions between helminthes and the host have been studied there is no research on the interaction between helminthes and the host?s gut microbiota and the impact on the host of worm-induced changes in its gut microbiota. Considering the importance of the gut microbiota to the health of the animal, this represents a new and exciting research area that we propose to address. Understanding this three-way interaction between the host, worm and microbiota that will have serious implications on our understanding of the pathology of parasitic infections The human whipworm (Trichuris trichiura) infects 1 billion people worldwide and resides in the large intestine, home to a complex diverse microbiota. Human whipworm infection has been successfully modeled using the mouse whip worm Trichuris muris and has generated many of the current paradigms of immunity to intestinal nematode infection. Critically, using this model the response phenotype to T. muris can easily be manipulated, thereby providing a setting whereby resistance or chronic infection can readily be investigated. Having demonstrated that chronic T. muris infection causes changes the mouse microbiota we now propose to use this model to study in detail the effect of a chronic infection on the host microbiota. The research objectives are: 1. To use metagenomics to reveal the changes in the mouse caecal microbiome during the establishment of a chronic T. muris infection. 2. To determine if a functional consequence of changes in the caecal microbiome associated with chronicity is an alteration in the ability of the microbiome to process complex dietary molecules, having long-term nutritional implications for chronically infected individuals. 3. To determine if a functional consequence of changes in the caecal microbiome associated with chronicity is to establish an ecosystem that influences subsequent T. muris egg hatching either decreasing it to reduce inter parasite competition or enhancing it thus allowing greater worm numbers to establish. 4. To demonstrate that changes in the microbiome will influence host immune status, are of a consequence of both the worm and the host response to it and are not maintained in the absence of the worm. Comprehension of the interactions between the host, worm and microbiota will increase our understanding of the pathology of parasitic infections, informing the way we might treat such infections.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION
Title: Simultaneous parenteral and pulmonary immunisation against tuberculosis
Funder: Medical Research Council

Abstract: Tuberculosis vaccinology is dominated by the prime boost paradigm. In this project we will establish the novel concept of simultaneous immunisation (SIM) against Mycobacterium tuberculosis (Mtb). We will determine the roles of different lung and peripheral lymphocyte populations in protective immunity induced by SIM in mice, and develop assays for protective pulmonary immunity in man. In mice intranasal immunisation with the subunit Mtb vaccine, Ad85A, generates pulmonary immunity. Importantly, Mtb growth is inhibited immediately after challenge. In contrast, parenteral BCG or subunit vaccines only inhibit Mtb growth late after challenge. The additive effect of parenteral/pulmonary immunisation does not depend on prime/boosting but on targeting early and late Mtb growth. Therefore SIM should be possible. We have demonstrated in practice that it is highly effective. We will determine the efficacy, duration and mechanism of protection of SIM. SIM safety will be tested by assessing immune responses and pathology after coincident infection with influenza virus or induction of allergic lung responses. Expression of the chemokine/receptor pair CXCL16/CXCR6 in the lungs after pulmonary immunisation is a correlate of protective pulmonary immunity. CXCR6+ cells are recoverable in bronchoalveolar lavage (BAL) unlike antigen specific cells induced by parenteral vaccines. However, three cell populations take part in pulmonary protection against Mtb, BAL cells, interstitial lung cells and extrapulmonary lymphocytes, but their roles in early and late inhibition of Mtb growth after challenge are ill-defined, as is the function of CXCL16/CXCR6 in intrapulmonary localization of cells. We shall use in vivo antibody depletion/blocking or transfer of purified cells to determine the function of different cells in early or late growth inhibition and the migration inhibitor, fingolimod, to dissect the contribution of intrapulmonary and extrapulmonary cells. CXCL16/CXCR6 deficient mice will illuminate the role of these molecules. We will develop assays for human pulmonary immunity by quantifying immune mediators in nasal and bronchial fluid or supernatants of cultured nasal epithelial cells, BAL or peripheral blood lymphocytes from Mtb un-exposed, Mtb exposed, latently infected subjects or Tb patients. This project will establish SIM as a highly effecive and safe method, which would require only a single clinic visit. We will determine the roles of different Tb-immune populations in protective lung immunity and the roles of CXCL16/CXCR6 in localization of cells within the lung. We will identify candidate assays for future assessment of immunity after pulmonary immunisation in man.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;3.4 VACCINES
Health categories: INFECTION
Title: Developmental Clinical Studies - Clinical evaluation of an AdCh63-MVA PvDBP_RII vaccine for blood-stage Plasmodium vivax
Funder: Medical Research Council

Abstract: The Need: Plasmodium vivax, one of the five Plasmodium species to cause human malaria, is the most widespread geographically and has been viewed, most certainly incorrectly, as a ?benign? parasite. In fact over 80 million cases of P. vivax malaria are reported annually, leading to severe morbidity for inhabitants of endemic areas as well as some mortality. Rationale: In recent years, viral vectored vaccines have been developed targeting the P. falciparum blood-stage malaria antigens MSP1 and AMA1. In Phase I/IIa clinical trials these vectors have induced strong serum antibody responses, unprecedented levels of cellular immunity and the first demonstration of efficacy with these antigens, albeit modest, in a human sporozoite challenge study. Unlike P. falciparum which utilises multiple redundant invasion pathways for human erythrocyte invasion, P. vivax requires an interaction with the Duffy blood group antigen, which is mediated by region II of the P. vivax Duffy-binding protein (PvDBP_RII). Given the essential nature of this invasion pathway, this candidate P. vivax antigen stands a far greater chance of success in comparison to the widely tested merozoite targets of P. falciparum. Solution & Development Plan: We have recently developed AdCh63 and MVA vectors expressing PvDBP_RII, and have shown in pre-clinical animal studies that, like for PfMSP1 and PfAMA1, these vectors induce strong antibody and cellular immunogenicity. This proposal aims to manufacture these two vectors to GMP and undertake a Phase Ia clinical trial to assess the safety, immunogenicity and in vitro efficacy of these new candidate vaccines for blood-stage P. vivax. If successful, these vaccines would be lead candidates for future efficacy testing.


Research activity codes: 6.1 PHARMACEUTICALS;3.4 VACCINES
Health categories: INFECTION
Title: 1. Identification of bacterial genes involved in antibiotic resistance using whole genome screens
Funder: Medical Research Council

Abstract: Antibiotic resistance in bacterial pathogens has spread, resulting in therapeutic failures in the treatment of some bacterial infections. A number of antibiotic resistance mechanisms are known, however, resistance is dependent on other, complementing mechanisms that have been termed ?innate resistance,? such as efflux systems which eject antibiotics from the bacterial cell. Recent work, and our proof of principle study, has identified genes not previously known to contribute to antibiotic resistance, and has indicated that other unknown resistance mechanisms exist. These numerous resistance mechanisms have been termed the bacterial ?resistome.? Transposon Directed Insertion-site Sequencing (TraDIS) combines high through-put Illumina sequencing and very large pools of transposon mutant libraries to simultaneously identify the site of every transposon insertion. This firstly identifies essential gene candidates, representing approximately 10% of the genome, in which transposon mutations are absent. These essential genes indicate candidate targets for the development of new antibacterial agents. Secondly, TraDIS allows every non-essential gene, approximately 90% of the genome, to be assayed simultaneously, which has not been achievable previously. Thus, by growing the mutant pool in sub-lethal concentrations of an antibiotic, the relative contribution of each non-essential gene to tolerance or sensitivity to the antibiotic may be measured. The entire non-essential gene set may then be listed in order of genes most important for resistance, through genes of no importance to those genes whose functions contribute to sensitivity. It is proposed to perform this type of experiment using two serotypes of Salmonella for which large transposon mutant libraries are already available; Typhi, which causes typhoid fever, and Typhimurium which causes food poisoning. In addition, transposon mutant collections will be generated for a methicillin resistant Staphylococcus aureus, an E. coli strain from a case of septicaemia, and Enterobacter cloacae, all of which cause nosocomial infections, and are particularly dangerous for immunocompromised and intensive care patients. A number of different antibiotics will be tested. This potentially provides new ways of combating resistance: resistant strains may be rendered sensitive using compounds that inhibit targets involved in resistance, and by compounds that induce expression of those genes products that contribute to sensitivity. In addition, the data will provide clues as to which resistance mechanisms may be operating in strains where the mechanism has not been identified. All the data will be made available on the Sanger Institute web site and will provide a useful resource for researchers engaged in antibiotic research and development.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: Structural and functional studies of Lassa fever virus multifunctional nucleoprotein
Funder: Medical Research Council

Abstract: Arenaviruses distribute worldwide, and many of them are important human pathogens. Particularly, Lassa fever viruses (LAVS) infect around 300,000 people and cause more than 5,000 deaths every year in South Africa. As no vaccine and effective drugs are available, LAVS is classed as a bioterrial reagent. The nucleoprotein (NP) of arenaviruses plays essential roles in immune evasion, viral RNA transcription, replication, RNA encapsdation and viral assembly, in which the mechanisms are unique and poorly understood. Following our most recently Nature article, we will use approaches of molecular virology, protein crystallography, Biochemistry, Biophysics and cell biology to reveal the detail mechanisms by testing our three hypotheses as below. Immune evasion: We hypothesize that the NP can specifically degrade the pathogen-associated pattern RNA ligands, so the cellular receptors RIG-I and MDA-5 can not detect the RNA ligands and not trigger the interferon pathways.We will determine how the C domain of the NP specifically recognize the pathogen-associated RNA ligands, and we will determine the structure of the NP C domain in complex with the RNA ligands. Cap-snatching: We hypothesize that the NP hides the cap (m7GpppG) of the cellular mRNAs and exposes the rest of the mRNAs that is been cleaved by endonuclease of the N-terminal domain of polymerase(L) and the trimmed by the 3 -5 nuclease of the NP. we will show exactly how the N-terminal domain of the NP binds the cap (m7-GpppG) by determining the complex structure and in vitro assays of the native and mutant proteins. We will try to form the RNA transcription and replication complexes (NP and RNA dependent polymerase L complex) in vitro for assays, and also check whether the complex can form short template RNA (1-4 nt). We will also try whether the N-terminal domain of the L protein can form complex with the NP, and try to crystallize the complex. NP multimerization and RNA encapisdation: We hypothesize that LAVS NP N-terminal residues and C-terminal residues are responsible for multimerization, and a highly charged clef of the NP is the viral RNA binding site. We will generate mutants to confirm the RNA binding cleft, and we will check the residues that are important for multimerization by mutagenesis, ITC and gel filtration. We will try to determine the NP-RNA complex structure by removing the E.coli RNAs and incubating with synthetic RNAs.In addition, We will study how the LAVS Z interacts with NP and L proteins .


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION
Title: Malaria parasite population structure and adaptation on the edge of endemic distribution in Africa
Funder: Medical Research Council

Abstract: Appropriate development of antimalarial vaccines and drugs needs to be informed by an understanding of processes of adaptation of malaria parasites in endemic populations. We will investigate population genetic structure and phenotypes of P. falciparum under selection at the north western extreme of its African range in Mauritania where endemicity persists despite limited and highly seasonal rainfall that restricts transmission. We predict that, compared with countries to the south that we recently studied, genetic divergence between populations of P. falciparum in different areas of Mauritania will be high due to genetic drift with small effective population sizes, and restricted gene flow among endemic foci. This will be investigated by conventional microsatellite analyses and whole genome sequence polymorphism analyses. We also predict that the selective landscape will be strikingly different from elsewhere, leading to measurable effects including the following: (i) a low level of acquired immunity in the human population will reduce diversifying selection on merozoite invasion ligands so these will be less diverse and highly skewed towards the use of a particular preferred primary ligand, which we can test with precision using a new erythrocyte culture and receptor knockdown system; (ii) alleles under strong positive selection will increase to high frequencies in local populations before sporadic gene flow events allow them to spread so there will be major local differences in drug resistance. Understanding these processes in terms of demographic and selective constraints on the parasite will guide targeting of existing control tools including antimalarial drugs and development of new tools including vaccines.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: Team CanUK: Novel antibacterial targets, assays, probes and opportunities in bacterial cell wall biogenesis
Funder: Medical Research Council

Abstract: There is a currently a global health threat posed by emerging antibiotic resistance amongst clinically important bacteria which is compounded by the decline in new drugs coming to the market. The bacterial cell wall (peptidoglycan, PG) is a validated, important, target for existing antibiotics and is a fundamental biological process, which is still poorly understood. Previous MRC funding in the UK has been used to create a research network focussed on PG with a synthesis facility at Warwick. This facility provides bespoke PG reagents to the research community that are not available commercially and has been a significant breakthrough in our capacity to address unanswered questions within the pathway. Researchers in Canada also have a track record of research in this area along with facilities for high throughput screening and compound libraries that can take advantage of new assays and substrates. The purpose of this UK-Canada network is thus to facilitate the development of bilateral research programmes with the aim of unravelling key features of bacterial cell wall biosynthesis using an integrated approach in which cell wall are analysed in a hierarchical manner from the fundamental biochemistry of peptidoglycan biochemistry, through sub-cellular organisation of the assembly machinery, to cell polymerisation. The aims are to understand the interrelationships between the formation of precursors, their export, polymerisation, modification and recycling to inform our fundamental understanding of the system. Overarching this is a desire to use this information in a rational manner to ultimately develop novel antimicrobials and identify new antimicrobial targets.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION