NIHRDataInsights/HRCSData huggingface dataset:
Title: Determining the importance of different locations to Mycobacterium tuberculosis transmission in high tuberculosis burden settings
Funder: Medical Research Council

Abstract: Currently, little is known about where M.tb. transmission occurs in high incidence settings, limiting the potential effectiveness of infection control interventions. The aim of this study is to estimate the proportion of transmission that occurs in different locations in Cape Town and other high incidence, sub-Saharan African settings, and to investigate the implications for TB control. There are four objectives: 1) Use network modelling to determine the effect of variation in infectiousness and susceptibility on the contribution of different location types to transmission. 2) Conduct primary data collection and analysis to inform model parameterisation. This objective has three parts: A. Estimate numbers of unique and repeat contacts by location (primary data collection using a questionnaire). B. Determine the suitability of different types of location for M.tb. transmission (data collection in buildings, including measuring CO2 and UV levels). C. Determine how contacts vary between people with different levels of behavioural and biological susceptibility to M.tb. infection and progression, by location (analysis of existing data). 3) Develop an individual-based model of contact behaviour and M.tb. transmission, and estimate the proportion of transmission that occurs in different locations in Cape Town. 4) Generalise the model to other settings, and investigate the impact of location-based infection control measures on M.tb. incidence and mortality. This study will directly address two of the WHO's research priorities: determining effects of infection control in 1) healthcare facilities and 2) other settings. Our findings will be of interest to international policy makers, and our results will be useful in informing and improving the effectiveness of infection control interventions. This understanding should contribute to meeting the 2025 WHO End TB Strategy Targets, in the high TB burden settings most in need of novel and innovative prevention strategies.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: A comparative trial of seasonal vaccination with the malaria vaccine RTS,S/AS01, seasonal malaria chemoprevention and the two interventions combined
Funder: Medical Research Council

Abstract: Methods for identifying children and ensuring that they receive the correct intervention, detecting clinical cases of malaria, determining the presence of anaemia and malnutrition and measuring severe adverse events have been developed during for the on-going SMC trial in the proposed study areas and appropriate standard operating procedures have been developed and are in place. The teams in Burkina Faso and Mali have each conducted successful malaria vaccine trials in the past so this component of the study should not pose any major technical issues. Measurement of anti-circumsporozoite (CSP) antibody concentrations induced by the vaccine, in particular the response to the fourth and fifth doses and the comparison in titres between children who receive a full or fractionated dose, will be important objectives of the trial as it is possible that repeated doses could lead to a diminishing response. To allow comparison of the results obtained in the new study with those of previous trials of RTS,S/AS01 sponsored by GSK, anti-CSP antibody measurements will be undertaken by the same contractor (The University of Ghent). The reason why the fourth dose of RTS,S/AS01 did not produce the kind of booster effect seen with many other vaccines is uncertain and, if repeated doses result in a progressive decline in antibody response, this would rule out seasonal vaccination. This study would provide the background for more detailed immunological investigation of this phenomenon, including its potential modulation, studies which might include measurement of antibody affinity, B cell function and memory and T cell immune responses. A request for support for these studies has not been included in this proposal but, if it is successful, additional funds will be sought to support more detailed immunological studies in partnership with a group with expertise in B cell immunology; one such group has already shown an interest in this proposal.


Research activity codes: 3.4 VACCINES
Health categories: INFECTION
Title: Assessing the safety of low dose primaquine in Plasmodium falciparum infected African children with glucose 6 phosphate dehydrogenase deficiency.
Funder: Medical Research Council

Abstract: Adding a single, low dose of primaquine (SLDPQ), 0.25 mg/kg, to the standard treatment of falciparum malaria is recommended by WHO for blocking malaria transmission. However countries are reluctant to use SLDPQ because of concerns that in glucose-6-phosphate deficient (G6PDd) individuals it may cause acute haemolytic anaemia, a well-known side effect of primaquine when given in higher, multiple, doses for vivax radical cure. Our study examines SDLPQ safety in children with acute uncomplicated falciparum malaria in a randomised, double blind, parallel group treatment trial of artemether-lumefantrine (AL)+SLDPQ vs. AL alone in G6PDd and in normal patients. We hypothesise that, compared to AL alone, SLDPQ+AL will not result in more profound anaemia (haemoglobin (Hb) <4 g/dL) or severe anaemia (Hb <5 g/dL) with features of severe malaria. The sample size is 1,572, 393/arm (80% power, one sided alpha 0.025), assuming a 1.5% event rate in G6PDd AL recipients and a 2.5% non-inferiority margin. Children from the Democratic Republic of Congo (Kinshasa) and Uganda (Soroti and Mbale) aged 6m-12y with Hb concentrations >= 6 g/dL will be recruited. Treated initially in hospital, they will be followed up daily for the first 5 days, day 7, then weekly to Day 42. Investigations include G6PD genotype & enzyme activity, malaria films (asexual and gametocyte counts), Hbs, molecular quantification of gametocytes, and primaquine and lumefantrine pharmacokinetics. Data analysis includes comparing rates of profound and severe anaemia, gametocyte clearance times, assessing risk factors for profound/severe anaemia, and modelling clinical, parasitological and pharmacokinetic factors for Hb recovery and gametocyte clearance. The results will provide high quality evidence on the safety of SLDPQ, informing drug policy for malaria elimination. They will also enable us to develop an optimised, age-based, SLDPQ regimen and so increase access to this valuable tool.


Research activity codes: 6.1 PHARMACEUTICALS
Health categories: INFECTION
Title: Global mechanisms for control of the trypanosome proteome: Defining the composition, origins and roles of cullin E3 ligases.
Funder: Medical Research Council

Abstract: Trypanosomatids evade the host immune response by multiple mechanisms, but for all species the parasite surface plays roles that are central to virulence. In Leishmania and the American trypanosome, surface-based mechanisms are focused on invasion and survival within host cells. By contrast, for African trypanosomes, where the parasite resides in the host bloodstream, the major mechanism is antigenic variation, but mounting evidence suggests that non-variant epitopes can be exposed to the immune system at the parasite surface. Some years ago we and others demonstrated that surface antibody is endocytosed and rapidly degraded, suggesting this as an additional component to antigenic variation in immune evasion. Furthermore, we also found that the major invariant surface glycoproteins (ISGs) were degraded by a ubiquitin-dependent reaction. However, we were unable to identify the proteins responsible for ubiquitylation of ISGs and hence could not advance our characterisation of the system. Significantly, ISGs are clearly immunogenic and are recognised by both human and bovine antibodies in serum from infected individuals. Recent work implicated cullin-class ubiquitin ligases as at least some of these ISG-interacting factors. Evidence comes from two major findings. Firstly, Usp7 and Vdu1, two deubiquitylating enzymes of ISGs, and which interact in mammalian cells with cullin ligases, modulate ISG expression levels. Secondly, knockdown of Skp1, a cullin adaptor recognising substrate proteins, is also involved in control of ISG expression levels. We propose to characterise cullin ligases in trypanosomes, to identify those that are responsible for surface protein turnover and to assess their overall roles in regulating protein turnover in trypanosomes. Given that cullin ligases are currently being exposed as possible drug targets, these studies will also contribute to assessing if cullins can be therapeutically exploited in trypanosomes.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: Membrane and host cytoskeleton reorganization during malaria parasite egress from erythrocytes
Funder: Medical Research Council

Abstract: Intracellular pathogens such as the malaria parasite hijack host cells and redirect their activity into facilitating replication and dispersal of the pathogen. The major clinically relevant phase of malaria is the asexual blood stage, during which parasites invade erythrocytes, multiply inside a vacuole and then escape for a new round of invasion. Egress of mature parasites from the erythrocyte requires the parasites to break through the two membranes enclosing them - the parasitophorous vacuole membrane and the erythrocyte cell membrane. These events are highly regulated, involving a cascade of activation of at least one parasite kinase, at least 2 proteases and likely other membrane modifying proteins such as phospholipases and/or pore-forming proteins. We have used electron tomography, video microscopy of live cells and soft X-ray tomography to uncover new details of the changes to the vacuolar and erythrocyte membranes and cytoskeleton during egress. These studies led to the discovery of a previously undescribed, initial step in egress, in which permeabilisation of the vacuole membrane precedes its complete disruption. This is rapidly followed by permeabilisation and breakage of the erythrocyte cell membrane and collapse of the cytoskeleton to allow the explosive release of parasites. With the methods we have developed, along with powerful new conditional gene modification tools in P. falciparum, we now wish to pursue the molecular and cellular basis of membrane and cytoskeleton reorganization during P. falciparum egress. Understanding the details of the egress process will lead to improved means of malaria control.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: G-quadruplex biology in the human malaria parasite Plasmodium falciparum
Funder: Medical Research Council

Abstract: The proposed work will investigate the distribution and metabolism of G-quadruplex (G4) motifs in both the DNA and RNA of the malaria parasite Plasmodium falciparum. G4-forming sequences are strikingly rare in the AT-rich P. falciparum genome, and are concentrated around the major family of virulence genes, called var genes. Our recent work has showed that G4 motifs may affect the expression and recombination of var genes, thus contributing to antigenic variation and diversification. Special classes of helicases are required to unwind G4s and we have knocked out two of these helicases in P. falciparum, revealing phenotypes in var gene expression, rates of genomic recombination, and telomere maintenance. We now propose to use genome-wide chromatin immunoprecipitation to establish the distribution of G4 motifs and their cognate helicases throughout the P. falciparum genome. At the RNA level, we will examine the G4 content of the transcriptome via RNA base modification and structure-specific sequencing. In a second arm of experiments, we will follow up our work on the two 'RecQ' G4 helicases by making and characterising a double RecQ knockout, and also a knockout of the second class of putative G4 helicase termed FANCJ. Finally, we will characterise these knockouts not only in terms of G4-specific phenotypes, but also in their effects on DNA replication dynamics throughout the genome, using DNA fibre analysis. Together, these experiments will provide multiple lines of evidence for the presence and roles of G4s and their cognate helicases in P. falciparum, thus improving our understanding of a novel aspect of malaria biology that is highly relevant to virulence.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: INFECTION
Title: Interaction and interdependency of GlmM and DacA - two essential enzymes required for methicillin resistance in Staphylococcus aureus
Funder: Medical Research Council

Abstract: The cell wall in Staphylococcus aureus is composed of a thick peptidoglycan layer, cell surface proteins, teichoic acids and carbohydrate polymers. Proper assembly of the different structures is not only essential for the pathogenesis of S. aureus but also for bacterial growth and several antimicrobial agents target the synthesis of key cell wall components. GlmM is a phosphoglucosamine mutase required for the production of UDP-N-acetylglucosamine, an essential cytoplasmic precursor for the synthesis of peptidoglycan. In S. aureus and many other Gram-positive bacteria, glmM forms a three-gene operon with dacA and ybbR, which encode the c-di-AMP synthase DacA and the membrane protein YbbR that is thought to regulate the activity of cylcase, respectively. c-di-AMP is a recently discovered signalling molecule that is essential for growth of S. aureus and many other Gram-positive bacteria in standard laboratory conditions. Recent work has highlighted a physical interaction between GlmM and DacA, suggesting that these enzymes are central players in coordinating peptidoglycan synthesis with the metabolic pathways that are regulated by c-di-AMP. These include in S. aureus potassium and osmolyte uptake as well as pH homeostasis. The aims of this project are to characterize the interdependency of the DacA and GlmM enzyme activities using in vitro assay systems, to determine the structure of the DacA/GlmM complex, to investigate the in vivo dynamics and the functional consequence of the DacA/GlmM interaction on the metabolism and methicillin resistance of S. aureus and to conclusively determine if GlmM is essential for the growth of S. aureus to resolve conflicting data. All together this work will allow us to propose a model for the molecular mechanism by which binding of GlmM can influence the activity of the c-di-AMP cyclase, provide insight into the biological significance of this protein interaction and uncover new avenues to interfere with the growth of S. aureus.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION
Title: Near-Infrared Spectroscopy: A One-Stop-Shop for Mosquito Epidemiological Monitoring?
Funder: Medical Research Council

Abstract: Mosquito control through the use of bednets, residual sprays or fumigation remains the most effective tool for preventing malaria and dengue though there is no easy to use entomological methods for directly assessing its efficacy. The rise of insecticide resistant mosquitoes exacerbates the need to understand how well interventions are working as it unclear whether control programmes should invest in more expensive insecticides. Mosquito age is the most informative method for evaluating the efficacy of adulticiding techniques though there are still no easy way of measuring it in the field. Near-Infrared Spectroscopy (NIRS) has been shown in the laboratory to be a promising, potentially high-throughput method that can both age mosquitoes and differentiate between sibling species though it has never been evaluated in the field. NIRS has relatively high individual mosquito measurement error though preliminary evidence suggests that precise mean estimates of the age of the population can be made by improving the machine learning methods converting spectral data into age and by fitting parametric models to groups of mosquitoes. This project proposes to rigorously test the ability of NIRS to monitor wild caught Anopheline and Aedes mosquito populations in semi-field and field sites. It shall use mathematical models to determine whether the mean age is sufficiently accurate to guide the use of vector control. The sampling requirements needed to estimate the effect size of an intervention will be defined and the methods tested in a malaria indoor-residual spraying programme in Africa. Evidence suggests that NIRS is also able to identify mosquitoes infected with dengue virus. This work will be extended to malaria to determine whether NIRS can differentiate uninfected, infected and infectious mosquitoes. Overall the proposal aims to generate the body of evidence needed to determine whether NIRS can be used to routinely monitor mosquito population in the field.


Research activity codes: 3.2 INTERVENTIONS TO ALTER PHYSICAL AND BIOLOGICAL ENVIRONMENTAL RISKS
Health categories: INFECTION
Title: VACCINE: Development and evaluation of vectored vaccines for HCV using an enhanced gene expression technology in a novel rodent hepacivirus model
Funder: Medical Research Council

Abstract: This project comprises the construction of a new type of vaccine vectored by cytomegalovirus (CMV) for immunisation against a rodent homologue of hepatitis C virus and its evaluation in a rodent / virus model along with an adenovirus (AdV) construct of previously demonstrated efficacy in animal models. Rodent hepacivirus (RHV) recapitulates in rats the liver-specific pathology, immune response and persistence observed in HCV infections in humans. Vectored constructs will be engineered to express transgenes of RHV and evaluated for efficacy as both a protective and therapeutic vaccine in a small animal model. The infectivity and immune responses to CMV and AdV-vectors in rats will be evaluated. Levels of RHV antigen expression will be determined and the effectiveness of a novel strategy to enhance gene expression through compositional alterations in transgene sequences investigated. We previously showed that frequencies of CpG and UpA dinucleotides in RNA viruses limit virus replication and gene expression; CMV and AdV both show evidence for similar host-imposed restrictions on expression that can be circumvented using synthetic transgenes with minimised CpG/UpA frequencies. The strength, breadth and durability of T cell responses to RHV and titres of neutralising antibodies in rats infected with vectors expressing compositionally altered constructs will be determined. In the 2nd phase of the project, CMV and AdV constructs will be evaluated for efficacy as a prophylactic vaccine in rat challenge experiments. As a therapeutic vaccine, RHV clearance will be monitored in RHV persistently infected rates post immunisation. Demonstration of efficacy of vaccines in this rodent model and determination of immune correlates of protection would translate directly into the future development and clinical trials of human vaccines for HCV.


Research activity codes: 3.4 VACCINES
Health categories: INFECTION
Title: T cells in tuberculosis: What type of T cell mediates immunity and how do we induce it by vaccination?
Funder: Medical Research Council

Abstract: TB represents a major challenge in vaccine development because we do not know constitutes a protective T cell and this is a complex issue in TB as unrestrained T cell responses are pathogenic and T cells must function for prolonged periods in inflammatory lesions to maintain long-term control. We propose that in TB, inflammation limits T cell function in ways that we do not fully understand and that we do not know how to prime T cells that function optimally in inflamed TB lesions, nor do we know what factors limit efficacy of T cells in these lesions. Importantly, we and others have recently determined that antigen-specific CD4 T cells within Mtb lesions exist in novel phenotypic subsets; some express the surface markers PD1 and CD69 and others express the marker KLRG1. The PD-1posCD69pos cells produce IL-2, express the IL-2 and IL-7 receptors, require ICOS and the transcription factor Bcl6 and locate within the parenchyma whereas the KLRG1pos cells express very high levels of the transcription factor T-bet, require IL-27R expression, produce the cytokine IFN and locate within the vasculature. Importantly, when transferred to an Mtb-infected lung, the PD-1posCD69pos population persist while the KLRG1pos population do not and the PD-1posCD69pos CD4 T cells mediate improved bacterial control whereas the KLRG1pos CD4 T cells, while expressing high levels of IFN, do not. Our working model is that protective CD4 T cells penetrate and persist within the mycobacterial lesions but that many vaccine-induced CD4 T cells fail to do this. We hypothesize that TCR-mediated events during priming define the function of T cells and that manipulation of the mycobacterial lesion will permit optimal function of T cells. We will develop a working model of how to generate T cells that function in the context of mycobacterial lesions, which will underpin rational design of vaccine strategies, development of potential correlates of protection and development of host directed therapies.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFECTION
Title: Novel strategies to detect and eliminate persistent Mycobacterium tuberculosis - applications in a murine Cornell model
Funder: Medical Research Council

Abstract: 1.Treatment with novel regimens in a modified Cornell model. The novel drug regimens containing rifapentine, bedaquiline, moxifloxacin will be tested in a modified Cornell mouse model in which treatment starts at 10 weeks after a low dose M. tuberculosis H37Rv infection. Treatment is given for 16 weeks by daily oral administration. Mouse organs will be harvested at two weeks interval to monitor CFU and broth counts (7H9 and culture filtrates). After treatment, the remaining mice are administered hydrocortisone for 8 week. CFU counts from lungs and spleens are performed to determine relapse. 2.Standardizing resuscitation promoting factor production and preparation of culture filtrates. M. tuberculosis is grown in 7H9 for 15 to 20 days until an optical density of 1 to 1.5 is reached. mRNA expression of 5 RPFs from the cultures is examined by semi-quantitative RT-PCR. The cultures are harvested by centrifugation and filtered with 0.2 um filter twice. The culture filtrates will be used immediately and RPF stability testing will be performed before and after storage at -20 or -70oC. 3.Resuscitation of M. tuberculosis in mouse organs. Broth counting is performed as serial 10-fold dilutions in triplicate in which 0.5 ml of organ homogenates are added to 4.5 ml of culture filtrates or 7H9. At 10-day intervals over a 2-month period of incubation, the broth cultures are examined for visible turbidity and confirmed by colonial morphology on 7H11 agar. The most probable numbers of viable bacilli are estimated from the patterns of positive and negative tubes. 4.Resuscitation of RPF-dependent persisters in Mycobacteria Growth Indicator Tube (MGIT). CFU count negative organ homogenates are incubated in MGITs, then culture filtrate is added into MGIT before and after 42 day incubation and continue to incubate for up to 2 months. Resuscitated persisters in MGIT tubes will be confirmed by observation of turbidity and fluorescence changes as well as plating on 7H11 agar plates.


Research activity codes: 5.1 PHARMACEUTICALS
Health categories: INFECTION