Imperial College London

ProfessorRobertWilkinson

Faculty of MedicineDepartment of Infectious Disease

Professor in Infectious Diseases
 
 
 
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Contact

 

+44 (0)20 7594 3891r.j.wilkinson Website

 
 
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Location

 

Wright Fleming WingSt Mary's Campus

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Summary

 

Publications

Publication Type
Year
to

354 results found

Boyles T, Stadelman A, Ellis JP, Cresswell FV, Lutje V, Wasserman S, Tiffin N, Wilkinson Ret al., The diagnosis of tuberculous meningitis in adults and adolescents: protocol for a systematic review and individual patient data meta-analysis to inform a multivariable prediction model, Wellcome Open Research, Vol: 4, Pages: 19-19

<ns4:p><ns4:bold>Background: </ns4:bold>Tuberculous meningitis (TBM) is the most lethal and disabling form of tuberculosis. Delayed diagnosis and treatment, which is a risk factor for poor outcome, is caused in part by lack of availability of diagnostic tests that are both rapid and accurate. Several attempts have been made to develop clinical scoring systems to fill this gap, but none have performed sufficiently well to be broadly implemented. We aim to identify and validate a set of clinical predictors that accurately classify TBM using individual patient data (IPD) from published studies.</ns4:p><ns4:p> <ns4:bold>Methods: </ns4:bold>We will perform a systematic review and obtain IPD from studies published from the year 1990 which undertook diagnostic testing for TBM in adolescents or adults using at least one of, microscopy for acid-fast bacilli, commercial nucleic acid amplification test for <ns4:italic>Mycobacterium tuberculosis</ns4:italic> or mycobacterial culture of cerebrospinal fluid.  Clinical data that have previously been shown to be associated with TBM, and can inform the final diagnosis, will be requested. The data-set will be divided into training and test/validation data-sets for model building. A predictive logistic model will be built using a training set with patients with definite TBM and no TBM. Should it be warranted, factor analysis may be employed, depending on evidence for multicollinearity or the case for including latent variables in the model.</ns4:p><ns4:p> <ns4:bold>Discussion: </ns4:bold>We will systematically identify and extract key clinical parameters associated with TBM from published studies and use a ‘big data’ approach to develop and validate a clinical prediction model with enhanced generalisability. The final model will be made available through a smartphone application. Further work will be external validation of the model and test

Journal article

Kubjane M, Berkowitz N, Goliath RT, Levitt N, Wilkinson RJ, Oni Tet al., Tuberculosis, HIV and the association with transient hyperglycaemia in peri-urban South Africa, Clinical Infectious Diseases, ISSN: 1058-4838

BackgroundDiabetes mellitus (DM) increases tuberculosis (TB) risk. We assessed the prevalence of hyperglycaemia (DM and impaired glucose regulation (IGR)) in TB patients and the association between hyperglycaemia and TB at enrolment and 3 months after TB treatment in the context of HIV-infection.MethodsAdults presenting at a Cape Town TB clinic were enrolled. TB cases were defined by South African guidelines, while non-TB participants were those who presented with respiratory symptoms, negative TB tests and resolution of symptoms 3 months later without TB treatment. HIV status was ascertained through medical records or HIV-testing. All participants were screened for DM using HbA1c and fasting plasma glucose at TB treatment and after 3 months. The association between TB and DM was assessed.ResultsOverall DM prevalence was 11.9% (95% CI: 9.1–15.4) at enrolment and 9.3% (95% CI: 6.4–13) at follow-up; IGR prevalence was 46.9% (95% CI 42.2–51.8) and 21.5% (95% CI 16.9–26.3) at enrolment and follow-up. TB/DM association was significant at enrolment (OR 2.41 (95% CI 1.3–4.3)) and follow-up (OR 3.3 (95% CI 1.5–7.3)), whilst TB/IGR association was only positive at enrolment OR 2.3 (95% CI 1.6–3.3). The TB/DM association was significant at enrolment in both new and pre-existing DM, but only persisted at follow-up in HIV-1-infected pre-existing DM.ConclusionOur study demonstrated high prevalence of transient hyperglycaemia and a significant TB/DM and TB/IGR association at enrolment in newly diagnosed DM, but persistent hyperglycaemia and TB/DM association in HIV-1-infected pre-existing DM, despite TB therapy.

Journal article

Tait DR, Hatherill M, van der Meeren O, Ginsburg AM, Van Brakel E, Salaun B, Scriba TJ, Akite EJ, Ayles HM, Bollaerts A, Demoitié M-A, Diacon A, Evans TG, Gillard P, Hellstrӧm E, Innes JC, Lempicki M, Malahleha M, Martinson N, Vela DM, Muyoyeta M, Nduba V, Pascal TG, Tameris M, Thienemann F, Wilkinson RJ, Roman Fet al., Final Analysis of a Trial of M72/AS01E Vaccine to Prevent Tuberculosis, New England Journal of Medicine, ISSN: 0028-4793

Journal article

Rohlwink U, Figaji A, Wilkinson K, Horswell S, Sesay A, Deffur A, Enslin N, Solomons R, Van Toorn R, Eley B, Levin M, Wilkinson R, Lai PJet al., Tuberculous meningitis in children is characterized by compartmentalized immune responses and neural excitotoxicity, Nature Communications, ISSN: 2041-1723

Tuberculous meningitis (TBM) is the most severe form of TB with high rates of mortality and morbidity. We conducted RNA-sequencing on whole blood as well as on ventricular and lumbar cerebrospinal fluid (CSF) of pediatric patients treated for TBM. Differential transcript expression of TBM cases were compared with healthy controls in whole blood and with non-TB cerebral infection controls in CSF. Whole blood RNA-Seq analysis demonstratesa distinct immune response pattern in TBM, with significant increase inboth canonical and non-canonical inflammasome activation and decrease in T-cell activation. In ventricular CSF, a significant enrichment associated with neuronal excitotoxicity and cerebral damage is detected in TBM. Finally, compartmental comparison in TBM indicatesthat the ventricular profile representsbrain injury whereas the lumbar profile representsprotein translation and cytokine signalling. Together, transcriptomic analysis shows thatdisease processes differ between the periphery and the central nervous system, and within brain compartments.

Journal article

Riou CR, Jhilmeet N, Rangaka MX, Wilkinson RJ, Wilkinson KAet al., Tuberculosis antigen-specific T cell responses during the first 6 months of antiretroviral treatment, Journal of Infectious Diseases, ISSN: 0022-1899

The reconstitution of Mycobacterium tuberculosis (Mtb)-antigen-specific CD4 T cells in a cohort of HIV-infected persons starting antiretroviral treatment (ART) in high TB endemic area is described. Restoration of the antigen-specific CD4 T cell subsets mirrored the overall CD4 T cell compartment. Activation (assessed by HLA-DR expression) decreased during ART but remained elevated compared to HIV-uninfected persons. Despite known Mtb sensitisation determined by IGRA, 12/23 participants had no Mtb-specific CD4 T cells detectable by flow cytometry, combined with overall elevated T cell activation and memory differentiation, suggesting heightened turnover. Our data suggest early ART initiation to maintain polyfunctional immune memory responses.

Journal article

Bunjun R, Omondi FMA, Makatsa M, Müller T, Prentice CSL, Wilkinson R, Riou C, Burgers Wet al., 2019, Th22 cells are a major contributor to the mycobacterial CD4+ T cell response and are depleted during HIV infection, Publisher: bioRxiv

HIV-1 infection substantially increases the risk of developing tuberculosis (TB). Some mechanisms, such as defects in the Th1 response to Mycobacterium tuberculosis ( M.tb ) in HIV-infected individuals have been widely reported. However, Th1-independent mechanisms also contribute to protection against TB. To identify a broader spectrum of defects in TB immunity during HIV infection, we examined IL-17 and IL-22 production in response to mycobacterial antigens in individuals with latent TB infection (LTBI) and HIV co-infection. Upon stimulating with mycobacterial antigens, we observed a distinct CD4+ T helper lineage producing IL-22 in the absence of IL-17 and IFN-γ. Th22 cells were present at high frequencies in response to mycobacterial antigens in blood and contributed up to 50% to the CD4+ T cell response to mycobacteria, comparable in magnitude to the IFN-γ Th1 response (median 0.91% and 0.55%, respectively). Phenotypic characterization of Th22 cells revealed that their memory differentiation was similar to M.tb -specific Th1 cells ( i.e . predominantly early-differentiated CD45RO+CD27+ phenotype). Moreover, CCR6 and CXCR3 expression profiles of Th22 cells were similar to Th17 cells, while their CCR4 and CCR10 expression patterns displayed an intermediate phenotype between Th1 and Th17 cells. Strikingly, mycobacterial IL-22 responses were three-fold lower in HIV-infected individuals compared to uninfected individuals, and the magnitude of responses correlated inversely with HIV viral load. These data provide important insights into mycobacteria-specific T helper subsets and suggest a potential role for IL-22 in protection against TB during HIV infection. Further studies are needed to fully elucidate the role of IL-22 in protective TB immunity.

Working paper

Wasserman S, Davis A, Wilkinson RJ, Meintjes Get al., Key considerations in the pharmacotherapy of tuberculous meningitis., Expert Opinion on Pharmacotherapy, Pages: 1-5, ISSN: 1465-6566

Journal article

Lesosky M, Rangaka MX, Pienaar C, Coussens AK, Goliath RT, Mathee S, Mwansa-Kambafwile J, Maartens G, Wilkinson RJ, Wilkinson KAet al., 2019, Plasma Biomarkers to Detect Prevalent or Predict Progressive Tuberculosis Associated With Human Immunodeficiency Virus–1, Clinical Infectious Diseases, Vol: 69, Pages: 295-305, ISSN: 1058-4838

BackgroundThe risk of HIV-1 infected individuals developing TB is high while both prognostic and diagnostic tools remain insensitive. The predictive performance of plasma biomarkers to identify HIV-1 infected individuals likely to progress to active disease is unknown.MethodsThirteen preselected analytes were determined from QuantiFERON® Gold in-tube (QFT) plasma samples in 421 HIV-1 infected persons recruited within the screening and enrolment phases of a randomised controlled trial of isoniazid preventive therapy. Blood for QFT was obtained pre-randomisation. Individuals were classified into prevalent TB, incident TB and controls. Comparisons between groups, supervised learning methods and weighted correlation network analyses were applied utilising the unstimulated and background-corrected plasma analyte concentrations.ResultsUnstimulated samples showed higher analyte concentrations in prevalent and incident TB compared to controls. The largest differences were seen for CXCL10, IL-2, IL-1 and TGF-. Predictive model analysis using unstimulated analytes discriminated better between controls and prevalent TB (Area Under the Curve AUC= 0·9), reasonably between incident and prevalent TB (AUC > 0·8), but poorly between controls and incident TB. Unstimulated IL-2 and IFN-γ were ranked at or near the top for all comparisons except the comparison between controls vs incident TB. Models using background adjusted values performed poorly.ConclusionsSingle plasma biomarkers are unlikely to distinguish between disease states in HIV-1 co-infected individuals and combinations of biomarkers are required. The ability to detect prevalent TB is potentially important, as no blood test hitherto has suggested utility to detect prevalent TB amongst HIV-1 co-infected persons.

Journal article

Deffur A, Wilkinson R, Mayosi B, Mulder Net al., 2019, ANIMA: Association Network Integration for Multiscale Analysis, Wellcome Open Research, Vol: 3, Pages: 1-33, ISSN: 2398-502X

Contextual functional interpretation of -omics data derived from clinical samples is a classical and difficult problem in computational systems biology. The measurement of thousands of datapoints on single samples has become routine but relating ‘big data’ datasets to the complexities of human pathobiology is an area of ongoing research. Complicating this is the fact that many publically available datasets use bulk transcriptomics data from complex tissues like blood. The most prevalent analytic approaches derive molecular ‘signatures’ of disease states or apply modular analysis frameworks to the data. Here we show, using a network-based data integration method using clinical phenotype and microarray data as inputs, that we can reconstruct multiple features (or endophenotypes) of disease states at various scales of organization, from transcript abundance patterns of individual genes through co-expression patterns of groups of genes to patterns of cellular behavior in whole blood samples, both in single experiments as well as in a meta-analysis of multiple datasets.

Journal article

Schutz C, Barr D, Andrade BB, Shey M, Ward A, Janssen S, Burton R, Wilkinson KA, Sossen B, Fukutani KF, Nicol MP, Maartens G, Wilkinson RJ, Meintjes Get al., Clinical, microbiologic, and immunologic determinants of mortality in hospitalized patients with HIV-associated tuberculosis: a prospective cohort study, PLoS Medicine, ISSN: 1549-1277

Background: In high burden settings case fatality rates are reported to be between 11% and 32% in hospitalized patients with HIV-associated tuberculosis, yet the underlying causes of mortality remain poorly characterized. Understanding causes of mortality could inform development of novel management strategies to improve survival. We aimed to assess clinical and microbiologic determinants of mortality and to characterize the pathophysiological processes underlying death by evaluating host soluble inflammatory mediators and determined the relationship between these mediators and death as well as biomarkers of disseminated tuberculosis. Methods and Findings: Adult HIV-positive patients hospitalized with a new diagnosis of HIV-associated tuberculosis were enrolled in Cape Town between 2014-2016. Detailed tuberculosis diagnostic testing was performed. Biomarkers of tuberculosis dissemination and host soluble inflammatory mediators at baseline were assessed. Of 682 enrolled participants, 576 with tuberculosis (487/576, 84.5% microbiologically confirmed) were included in analyses. The median age was 37 years (IQR=31-43), 51.2% were female and the patients had advanced HIV with median CD4 count =58 cells/l (IQR= 21-120) and median HIV viral load=5.1 log10 copies/mL (IQR=3.3-5.7).Antituberculosis therapy was initiated in 566/576 (98.3%) and 487/576 (84.5%) started therapy within 48 hours of enrolment. Twelve-week mortality was 124/576 (21.5%) with 46/124 (37.1%) deaths occurring within 7 days of enrolment. Clinical and microbiologic determinants of mortality included disseminated tuberculosis (positive urine lipoarabinomannan, urine Xpert MTB/RIF or tuberculosis blood culture in 79.6% of deaths vs 60.7% of survivors, p=0.001), sepsis syndrome (high lactate in 50.8% of deaths vs 28.9% of survivors, p<0.001) and rifampicin resistant tuberculosis (16.9% of deaths vs 7.2% of survivors, p=0.002). Using non-supervised two-way hierarchical cluster and principal components analy

Journal article

Rockwood N, Lai RPJ, Seldon R, Young D, Wilkinson Ret al., 2019, Variation in pre-therapy levels of selected Mycobacterium tuberculosis transcripts insputumand their relationship with 2-month culture conversion, Wellcome Open Research, Vol: 106, Pages: 1-9, ISSN: 2398-502X

Background: The abundance of transcripts arising from Mycobacterium tuberculosis (MTB) in sputum pre-chemotherapy may enhance our understanding of factors influencing treatment response. We hypothesized that differences in the prevalence of pre-existing slowly metabolizing MTB in sputum may be partially responsible for differences in the rate of sputum clearance during treatment. Methods: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to characterize a selected limited transcription profile of MTB in sputum pre-chemotherapy and assess inter-individual variation. The difference in cycle threshold (Ct) per gene, normalized to 16S, between exponential/stationary phase culture and sputum was calculated and stratified by 2-month culture converter status. Results: HIV-1 uninfected patients with rifampicin-susceptible tuberculosis provided sputum pre-chemotherapy; 11 patients were negative for MTB culture after two months of therapy and 8 remained culture-positive. Increased icl1 and prpD and rpsN2:rpsN1 in sputum relative to culture suggested cholesterol utilization and a low-zinc environment respectively. Increased hspX and decreased atpA and nuoG relative to exponential culture suggested a slowly metabolizing subpopulation of MTB. While the the hspX hi atpA lo nuoG lo signal varied, we did not observe statistically significant enrichment of this phenotype in the non-converter population nor an association with MTB-lineage. Conclusion: Differential abundance of selected informative transcripts suggested a metabolically less-active subpopulation with a prevalence that varied between individual untreated patients.

Journal article

Kouchaki S, Yang Y, Walker TM, Sarah Walker A, Wilson DJ, Peto TEA, Crook DW, Clifton DAet al., 2019, Application of machine learning techniques to tuberculosis drug resistance analysis, Bioinformatics, Vol: 35, Pages: 2276-2282, ISSN: 1367-4803

Timely identification of Mycobacterium tuberculosis (MTB) resistance to existing drugs is vital to decrease mortality and prevent the amplification of existing antibiotic resistance. Machine learning methods have been widely applied for timely predicting resistance of MTB given a specific drug and identifying resistance markers. However, they have been not validated on a large cohort of MTB samples from multi-centers across the world in terms of resistance prediction and resistance marker identification. Several machine learning classifiers and linear dimension reduction techniques were developed and compared for a cohort of 13 402 isolates collected from 16 countries across 6 continents and tested 11 drugs.</jats:p> </jats:sec> <jats:sec> <jats:title>Results</jats:title> <jats:p>Compared to conventional molecular diagnostic test, area under curve of the best machine learning classifier increased for all drugs especially by 23.11%, 15.22% and 10.14% for pyrazinamide, ciprofloxacin and ofloxacin, respectively (P &lt; 0.01). Logistic regression and gradient tree boosting found to perform better than other techniques. Moreover, logistic regression/gradient tree boosting with a sparse principal component analysis/non-negative matrix factorization step compared with the classifier alone enhanced the best performance in terms of F1-score by 12.54%, 4.61%, 7.45% and 9.58% for amikacin, moxifloxacin, ofloxacin and capreomycin, respectively, as well increasing area under curve for amikacin and capreomycin. Results provided a comprehensive comparison of various techniques and confirmed the application of machine learning for better prediction of the large diverse tuberculosis data. Furthermore, mutation ranking showed the possibility of finding new resistance/susceptible markers.</jats:p> </jats:sec> <jats:sec>

Journal article

Walker NF, Opondo C, Meintjes GA, Jhilmeet N, Friedland J, Elkington PT, Wilkinson RJ, Wilkinson KAet al., 2019, Invariant Natural Killer T cell dynamics in HIV-associated tuberculosis, Clinical Infectious Diseases, ISSN: 1058-4838

RationaleTuberculosis (TB) is the leading cause of mortality and morbidity in people living with HIV infection. HIV-infected patients with TB disease are at risk of the paradoxical TB-associated immune reconstitution inflammatory syndrome (TB-IRIS) when they commence anti-retroviral therapy. However, the pathophysiology is incompletely understood and specific therapy is lacking.ObjectivesWe investigated the hypothesis that invariant Natural Killer T (iNKT) cells contribute to innate immune dysfunction associated with TB-IRIS.MethodsIn a cross-sectional study of 101 HIV-infected and -uninfected South African patients with active TB and controls, iNKT cells were enumerated using α-galactosylceramide-loaded CD1d tetramers and subsequently functionally characterised by flow cytometry. In a second study of 49 HIV-1-infected TB patients commencing anti-retroviral therapy, iNKT cells in TB-IRIS patients with non-IRIS controls were compared longitudinally.Measurements and main resultsCirculating iNKT cells were reduced in HIV-1 infection, most significantly the CD4+ subset, which was inversely associated with HIV-1 viral load. iNKT cells in HIV-associated TB had increased surface CD107a expression, indicating cytotoxic degranulation. Relatively increased iNKT cell frequency in HIV-infected patients with active TB was associated with development of TB-IRIS following anti-retroviral therapy initiation. iNKT cells in TB-IRIS were CD4+CD8- subset deplete and degranulated around the time of TB-IRIS onset.ConclusionsReduced iNKT cell CD4+ subsets as a result of HIV-1 infection may skew iNKT cell functionality towards cytotoxicity. Increased CD4- cytotoxic iNKT cells may contribute to immunopathology in TB-IRIS.

Journal article

Daskapan A, Idrus LR, Postma MJ, Wilffert B, Kosterink JGW, Stienstra Y, Touw DJ, Andersen AB, Bekker A, Denti P, Hemanth Kumar AK, Kidola J, Kwara A, McIlleron H, Meintjes G, van Oosterhout JJ, Ramachandran G, Rockwood NS, Wilkinson RJ, van der Werf TS, Alfefenar J-WCet al., 2019, A systematic review on the effect of HIV infection on the pharmacokinetics of first-line tuberculosis drugs, Clinical Pharmacokinetics, Vol: 58, Pages: 747-766, ISSN: 1179-1926

IntroductionContrasting findings have been published regarding the effect of human immunodeficiency virus (HIV) on tuberculosis (TB) drug pharmacokinetics (PK).ObjectivesThe aim of this systematic review was to investigate the effect of HIV infection on the PK of the first-line TB drugs (FLDs) rifampicin, isoniazid, pyrazinamide and ethambutol by assessing all published literature.MethodsSearches were performed in MEDLINE (through PubMed) and EMBASE to find original studies evaluating the effect of HIV infection on the PK of FLDs. The included studies were assessed for bias and clinical relevance. PK data were extracted to provide insight into the difference of FLD PK between HIV-positive and HIV-negative TB patients. This systematic review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement and its protocol was registered at PROSPERO (registration number CRD42017067250).ResultsOverall, 27 studies were eligible for inclusion. The available studies provide a heterogeneous dataset from which consistent results could not be obtained. In both HIV-positive and HIV-negative TB groups, rifampicin (13 of 15) and ethambutol (4 of 8) peak concentration (Cmax) often did not achieve the minimum reference values. More than half of the studies (11 of 20) that included both HIV-positive and HIV-negative TB groups showed statistically significantly altered FLD area under the concentration–time curve and/or Cmax for at least one FLD.ConclusionsHIV infection may be one of several factors that reduce FLD exposure. We could not make general recommendations with respect to the role of dosing. There is a need for consistent and homogeneous studies to be conducted.

Journal article

Gliddon HD, Kaforou M, Alikian M, Habgood-Coote D, Zhou C, Oni T, Anderson ST, Brent AJ, Crampin AC, Eley B, Kern F, Langford PR, Ottenhoff THM, Hibberd ML, French N, Wright VJ, Dockrell HM, Coin LJ, Wilkinson RJ, Levin Met al., 2019, Identification of reduced host transcriptomic signatures for tuberculosis and digital PCR-based validation and quantification, Publisher: Cold Spring Harbor Laboratory

<jats:title>Abstract</jats:title><jats:p>Recently, host whole blood gene expression signatures have been identified for diagnosis of tuberculosis (TB). Absolute quantification of the concentrations of signature transcripts in blood have not been reported, but would facilitate the development of diagnostic tests.</jats:p><jats:p>To identify minimal transcript signatures, we applied a novel transcript selection procedure to microarray data from African adults comprising 536 patients with TB, other diseases (OD) and latent TB (LTBI), divided into training and test sets. Signatures were validated using reverse transcriptase (RT) - digital PCR (dPCR).</jats:p><jats:p>A four-transcript signature (<jats:italic>GBP6</jats:italic>, <jats:italic>TMCC1</jats:italic>, <jats:italic>PRDM1</jats:italic>, <jats:italic>ARG1</jats:italic>) measured using RT-dPCR distinguished TB patients from those with OD (area under the curve (AUC) 93.8% (CI<jats:sub>95%</jats:sub> 82.2 – 100%). A three-transcript signature (<jats:italic>FCGR1A, ZNF296, C1QB</jats:italic>) differentiated TB from LTBI (AUC 97.3%, CI<jats:sub>95%</jats:sub>: 93.3 – 100%), regardless of HIV.</jats:p><jats:p>These signatures have been validated across platforms and across samples offering strong, quantitative support for their use as diagnostic biomarkers for TB.</jats:p>

Working paper

Rohlwink UK, Walker NF, Ordonez AA, Li YJ, Tucker EW, Elkington PT, Wilkinson RJ, Wilkinson KAet al., 2019, Matrix metalloproteinases in pulmonary and central nervous system tuberculosis-a review, International Journal of Molecular Sciences, Vol: 20, ISSN: 1422-0067

Tuberculosis (TB) remains the single biggest infectious cause of death globally, claiming almost two million lives and causing disease in over 10 million individuals annually. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes with various physiological roles implicated as key factors contributing to the spread of TB. They are involved in the breakdown of lung extracellular matrix and the consequent release of Mycobacterium tuberculosis bacilli into the airways. Evidence demonstrates that MMPs also play a role in central nervous system (CNS) tuberculosis, as they contribute to the breakdown of the blood brain barrier and are associated with poor outcome in adults with tuberculous meningitis (TBM). However, in pediatric TBM, data indicate that MMPs may play a role in both pathology and recovery of the developing brain. MMPs also have a significant role in HIV-TB-associated immune reconstitution inflammatory syndrome in the lungs and the brain, and their modulation offers potential novel therapeutic avenues. This is a review of recent research on MMPs in pulmonary and CNS TB in adults and children and in the context of co-infection with HIV. We summarize different methods of MMP investigation and discuss the translational implications of MMP inhibition to reduce immunopathology.

Journal article

Kendall EA, Azman AS, Maartens G, Boulle A, Wilkinson RJ, Dowdy DW, Rangaka MXet al., 2019, Projected population-wide impact of antiretroviral therapy-linked isoniazid preventive therapy in a high-burden setting, AIDS, Vol: 33, Pages: 525-536, ISSN: 0269-9370

OBJECTIVE: Both isoniazid preventive therapy (IPT) and antiretroviral therapy (ART) reduce tuberculosis risk in individuals living with HIV. We sought to estimate the broader, population-wide impact of providing a pragmatically-implemented 12-month IPT regimen to ART recipients in a high-burden community. DESIGN: Dynamic transmission model of a tuberculosis-HIV epidemic, calibrated to site-specific, historical epidemiologic and clinical trial data from Khayelitsha, South Africa. METHODS: We projected the five-year impact of delivering a 12-month IPT regimen community-wide to 85% of new ART initiators and 15%/year of those already on ART, accounting for IPT-attributable reductions in TB infection, progression, and transmission. We also evaluated scenarios of continuously-delivered IPT, ongoing ART scale-up, and lower tuberculosis incidence. RESULTS: Under historical (early 2010 s) ART coverage, this ART-linked IPT intervention prevented one tuberculosis case per 18 (95% credible interval [CrI] 11-29) people treated. It lowered tuberculosis incidence by a projected 23% (95%CrI 14-30%) among people receiving ART, and by 5.2% (95%CrI 2.9-8.7%) in the total population. Continuous IPT reduced the number needed to treat to prevent one case of tuberculosis to 10 (95%CrI 7-16), though it required 74% more person-years of therapy (95%CrI 64-94%) to prevent one TB case, relative to 12-month therapy. Under expanding ART coverage, the tuberculosis incidence reduction achieved by 12-month IPT grew to 7.6% (95%CrI 4.3-12.6%). Effect sizes were similar in a simulated setting of lower tuberculosis incidence. CONCLUSIONS: IPT in conjunction with ART reduces tuberculosis incidence among those who receive therapy and has additional impact on tuberculosis transmission in the population.

Journal article

Zürcher K, Ballif M, Fenner L, Borrell S, Keller PM, Gnokoro J, Marcy O, Yotebieng M, Diero L, Carter EJ, Rockwood N, Wilkinson RJ, Cox HS, Ezati N, Abimiku A, Collantes J, Avihingsanon A, Kawkitinarong K, Reinhard M, Hömke R, Huebner R, Gagneux S, Böttger E, Egger Met al., 2019, Drug susceptibility testing and mortality in patients treated for tuberculosis in high-burden countries: A multi-centre cohort study, Lancet Infectious Diseases, Vol: 19, Pages: 298-307, ISSN: 1473-3099

Background: Drug resistance is a challenge for the global control of tuberculosis. We examined mortality in tuberculosis patients from high-burden countries, according to concordance or discordance of results from drug susceptibility testing (DST) done locally and in a reference laboratory.Methods: We collected Mycobacterium tuberculosis isolates from adult patients in Côte d’Ivoire, Democratic Republic of the Congo, Kenya, Nigeria, South Africa, Peru, and Thailand, stratified by HIV status and tuberculosis drug resistance. Molecular or phenotypic drug susceptibility testing (DST) was done locally and at the Swiss tuberculosis reference laboratory. We examined mortality during treatment according to DST results and treatment adequacy in logistic regression models adjusting for sex, age, sputum microscopy and HIV status.Findings: 634 tuberculosis patients were included; median age was 33.2 years, 239 (37.7%) were female, 272 (42.9%) HIV-positive and 69 (10.9%) patients died. Based on the reference laboratory DST, 394 (62.2%) strains were pan-susceptible, 45 (7.1%) mono-resistant, 163 (25.7%) multidrug-resistant (MDR-TB), and 30 (4.7%) had preextensive or extensive drug resistance (pre-XDR/XDR-TB). Results of reference and local laboratories were discordant in 121 (19.1%) cases. Overall, sensitivity and specificity to detect any resistance were 90.8% and 84.3%, respectively. Mortalityranged from 6.0% (20/336) in patients with pan-susceptible tuberculosis treated according to WHO guidelines to 57.1% (8/14) in patients with resistant strains who were under treated. In logistic regression, compared to concordant DST results, the adjusted odds ratio of death was 7.33 (95% CI 2.70-19.95) for patients with discordant results potentially leading to under treatment. Interpretation: Inaccurate DST by comparison to a reference standard led to under treatment of drug resistant tuberculosis and increased mortality. Rapid molecular DST of first- and second-line drugs a

Journal article

Li Y, Wilkinson KA, Wilkinson R, Figaji A, Rohlwink UKet al., 2019, Elevated matrix metalloproteinases offer novel insight into their role in pediatric tuberculous meningitis, Journal of the Pediatric Infectious Diseases Society, ISSN: 2048-7193

We collected lumbar and ventricular cerebrospinal fluid and serum from 40 children treated for tuberculous meningitis and measured the concentrations of gelatinases and their inhibitors. The concentrations of matrix metalloproteinase 9 (MMP-9), MMP-2, tissue inhibitor of metalloproteinase 1 (TIMP-1), and TIMP-2 were significantly elevated in the lumbar CSF samples, and we found interesting dynamics for MMP-9 that offer novel insight into its role in pediatric patients with tuberculous meningitis.

Journal article

Davis AG, Rohlwink UK, Proust A, Figaji A, Wilkinson RJet al., 2019, The pathogenesis of tuberculous meningitis, Journal of Leukocyte Biology, Vol: 105, Pages: 267-280, ISSN: 0741-5400

Tuberculosis (TB) remains a leading cause of death globally. Dissemination of TB to the brain results in the most severe form of extrapulmonary TB, tuberculous meningitis (TBM), which represents a medical emergency associated with high rates of mortality and disability. Via various mechanisms the Mycobacterium tuberculosis (M.tb) bacillus disseminates from the primary site of infection and overcomes protective barriers to enter the CNS. There it induces an inflammatory response involving both the peripheral and resident immune cells, which initiates a cascade of pathologic mechanisms that may either contain the disease or result in significant brain injury. Here we review the steps from primary infection to cerebral disease, factors that contribute to the virulence of the organism and the vulnerability of the host and discuss the immune response and the clinical manifestations arising. Priorities for future research directions are suggested.

Journal article

Boyles T, Stadelman A, Ellis J, Cresswell F, Lutje V, Wasserman S, Tiffin N, Wilkinson Ret al., 2019, The diagnosis of tuberculous meningitis in adults and adolescents: protocol for a systematic review and individual patient data meta-analysis to inform a multivariable prediction model, Wellcome Open Research, Vol: 4, ISSN: 2398-502X

Background: Tuberculous meningitis (TBM) is the most lethal and disabling form of tuberculosis. Delayed diagnosis and treatment, which is a risk factor for poor outcome, is caused in part by lack of availability of diagnostic tests that are both rapid and accurate. Several attempts have been made to develop clinical scoring systems to fill this gap, but none have performed sufficiently well to be broadly implemented. We aim to identify and validate a set of clinical predictors that accurately classify TBM using individual patient data (IPD) from published studies. Methods: We will perform a systematic review and obtain IPD from studies published from the year 1990 which undertook diagnostic testing for TBM in adolescents or adults using at least one of, microscopy for acid-fast bacilli, commercial nucleic acid amplification test for Mycobacterium tuberculosis or mycobacterial culture of cerebrospinal fluid.  Clinical data that have previously been shown to be associated with TBM, and can inform the final diagnosis, will be requested. The data-set will be divided into training and test/validation data-sets for model building. A predictive logistic model will be built using a training set with patients with definite TBM and no TBM. Should it be warranted, factor analysis may be employed, depending on evidence for multicollinearity or the case for including latent variables in the model. Discussion: We will systematically identify and extract key clinical parameters associated with TBM from published studies and use a ‘big data’ approach to develop and validate a clinical prediction model with enhanced generalisability. The final model will be made available through a smartphone application. Further work will be external validation of the model and test of efficacy in a randomised controlled trial.

Journal article

Jhilmeet N, Lowe DM, Riou CR, Scriba TJ, Coussens AK, Goliath RT, Wilkinson RJ, Wilkinson KAet al., 2018, The effect of antiretroviral treatment on selected genes in whole blood from HIV-infected adults sensitised by Mycobacterium tuberculosis, PLoS ONE, Vol: 13, ISSN: 1932-6203

HIV-1 co-infection is a leading cause of susceptibility to tuberculosis (TB), with the risk of TB being increased at all stages of HIV-1 infection. Antiretroviral treatment (ART) is the most effective way to reduce the risk of TB in HIV-1 co-infected people. Studying protective, ART-induced, immune restoration in HIV-1 infected individuals sensitised by Mycobacterium tuberculosis (Mtb) can thus help identify mechanisms of protection against TB. In order to understand ART-mediated prevention of TB in HIV-1 infected adults, we investigated the expression of 30 genes in whole blood from HIV-1 infected patients during the first 6 months of ART-induced immune reconstitution. The 30 selected genes were previously described to be differentially expressed between sorted Mtb specific central and effector memory CD4 T cells. HIV-1 infected persons sensitised by Mtb were recruited in Khayelitsha, South Africa, when initiating ART. RNA was extracted from whole blood at initiation and 1, 3 and 6 months of ART. qRT-PCR was used to determine gene expression and three reference ‘housekeeping’ genes were used to calculate the fold change in the expression of each gene relative to day 0 of ART. Results were assessed longitudinally. We observed a decrease in the expression of a number of genes at 6 months of ART, reflecting a decrease in immune activation. However, following correction for multiple comparisons and increasing CD4 counts, only the decrease in CD27 gene expression remained statistically significant. While not statistically significant, a number of genes also showed increased expression at various timepoints, illustrating the broad regeneration of the T cell pool in HIV-1 infected adults on ART. Our findings generate hypotheses underlying ART- induced protective immune reconstitution and may pave the way for future studies to evaluate ART mediated prevention of TB in HIV-1 infected persons.

Journal article

Heemskerk AD, Donovan J, Anh Thu DD, Marais S, Chaidir L, Mong Dung VT, Centner CM, Ngoc Ha VT, Annisa J, Dian S, Bovijn L, Hoang Mai NT, Phu NH, Vinh Chau NV, Rizal Ganiem A, Van CT, Geskus RB, Thuong Thuong NT, Ruslami R, Meintjes G, van Crevel R, Wilkinson RJ, Thwaites GEet al., 2018, Improving the microbiological diagnosis of tuberculous meningitis: a prospective, international, multicentre comparison of conventional and modified Ziehl-Neelsen stain, GeneXpert, and culture of cerebrospinal fluid, Journal of Infection, Vol: 77, Pages: 509-515, ISSN: 0163-4453

ObjectivesTuberculous meningitis (TBM) is the severest form of tuberculosis, but current diagnostic tests are insensitive. Recent reports suggest simple modifications to conventional cerebrospinal fluid (CSF) Ziehl–Neelsen (ZN) staining may greatly improve sensitivity. We sought to define the performance of modified and conventional ZN stain for TBM diagnosis.MethodsIn hospitals in Vietnam, South Africa and Indonesia we conducted a prospective study of modified ZN with or without cytospin, conventional ZN smear, GeneXpert, and culture on CSF in adults with suspected TBM.ResultsA total of 618 individuals were enrolled across 3 sites. Compared with the TBM clinical diagnostic gold standard for research (definite probable or possible TBM), sensitivity of conventional ZN and modified ZN with cytospin were 33.9% and 34.5% respectively (p = 1.0 for the difference between tests), compared with culture 31.8% and Xpert 25.1%. Using culture as a reference, sensitivities of conventional ZN, modified ZN with cytospin, and Xpert were 66.4%, 67.5%, and 72.3%, respectively. Higher CSF volume and lactate, and lower CSF:blood glucose ratio were independently associated with microbiologically confirmed TBM.ConclusionsModified ZN stain does not improve diagnosis of TBM. Currently available tests are insensitive, but testing large CSF volumes improves performance. New diagnostic tests for TBM are urgently required.

Journal article

Meintjes G, Stek C, Bluementhal L, Thienemann F, Schutz C, Buyze J, Ravinetto R, van Loen H, Nair A, Jackson A, Colebunders R, Maartens G, Wilkinson RJ, Lynen Let al., 2018, Prednisone for prevention of paradoxical tuberculosis-associated IRIS, New England Journal of Medicine, Vol: 379, Pages: 1915-1925, ISSN: 0028-4793

Background: Early initiation of antiretroviral therapy (ART) in patients with tuberculosis reduces mortality in those with low CD4 counts, but increases the risk of paradoxical tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS). We determined whether prophylactic prednisone safely reduces the incidence of paradoxical TB-IRIS in patients at high risk. Methods: Randomized, double-blind, placebo-controlled trial of prednisone (40 mg/day for 14 days, then 20 mg/day for 14 days) started with ART in ART-naïve adults at high risk of TB-IRIS (within 30 days of antituberculosis treatment initiation and CD4 count ≤100 cells/μl). Primary endpoint was development of TB-IRIS within 12 weeks, adjudicated by an independent committee.Results: Among 240 participants median age was 36 (IQR=30-42), 60% male, and median CD4 49 cells/μl (IQR=24-86). 18 participants were lost to follow-up or withdrew. TB-IRIS was diagnosed in 56 in the placebo arm (46.7%) and 39 in the prednisone arm (32.5%) (relative risk (RR)=0.70 (95%CI=0.51-0.96); p=0.03). Open-label corticosteroids to treat TB-IRIS were prescribed to 34 of the placebo arm (28.3%) and 16 (13.3%) of the prednisone arm (RR=0.47 (95%CI=0.27-0.81)). 4 deaths occurred in the placebo arm; 5 in the prednisone arm (p=1.0). Severe infections (AIDS-defining or invasive bacterial) occurred in 18 in the placebo arm; 11 in the prednisone arm (p=0.23). One Kaposi’s sarcoma case occurred (placebo arm).Conclusions: Prednisone during the first 4 weeks of initation of ART for HIV-1 infection reduced TB associated IRIS without evidence for increased risk of severe infections or malignancy.

Journal article

Deffur A, Wilkinson R, Mayosi B, Mulder Net al., 2018, ANIMA: Association network integration for multiscale analysis [version 3; referees: 2 approved, 1 approved with reservations], Wellcome Open Research, Vol: 3, ISSN: 2398-502X

Contextual functional interpretation of -omics data derived from clinical samples is a classical and difficult problem in computational systems biology. The measurement of thousands of data points on single samples has become routine but relating ‘big data’ datasets to the complexities of human pathobiology is an area of ongoing research. Complicating this is the fact that many publicly available datasets use bulk transcriptomics data from complex tissues like blood. The most prevalent analytic approaches derive molecular ‘signatures’ of disease states or apply modular analysis frameworks to the data. Here we describe ANIMA (association network integration for multiscale analysis), a network-based data integration method using clinical phenotype and microarray data as inputs. ANIMA is implemented in R and Neo4j and runs in Docker containers. In short, the build algorithm iterates over one or more transcriptomics datasets to generate a large, multipartite association network by executing multiple independent analytic steps (differential expression, deconvolution, modular analysis based on co-expression, pathway analysis) and integrating the results. Once the network is built, it can be queried directly using Cypher (a graph query language), or by custom functions that communicate with the graph database via language-specific APIs. We developed a web application using Shiny, which provides fully interactive, multiscale views of the data. Using our approach, we show that we can reconstruct multiple features of disease states at various scales of organization, from transcript abundance patterns of individual genes through co-expression patterns of groups of genes to patterns of cellular behaviour in whole blood samples, both in single experiments as well in meta-analyses of multiple datasets.

Journal article

Kroon E, Coussens AK, Kinnear C, Orlova M, Muller M, Seeger A, Wilkinson RJ, Hoal E, Schurr Eet al., 2018, Neutrophils: innate effectors of TB resistance?, Frontiers in Immunology, Vol: 9, ISSN: 1664-3224

Certain individuals are able to resist Mycobacterium tuberculosis infection despite persistent and intense exposure. These persons do not exhibit adaptive immune priming as measured by tuberculin skin test (TST) and interferon-γ (IFN-γ) release assay (IGRA) responses, nor do they develop active tuberculosis (TB). Genetic investigation of individuals who are able to resist M. tuberculosis infection shows there are likely a combination of genetic variants that contribute to the phenotype. The contribution of the innate immune system and the exact cells involved in this phenotype remain incompletely elucidated. Neutrophils are prominent candidates for possible involvement as primers for microbial clearance. Significant variability is observed in neutrophil gene expression and DNA methylation. Furthermore, inter-individual variability is seen between the mycobactericidal capacities of donor neutrophils. Clearance of M. tuberculosis infection is favored by the mycobactericidal activity of neutrophils, apoptosis, effective clearance of cells by macrophages, and resolution of inflammation. In this review we will discuss the different mechanisms neutrophils utilize to clear M. tuberculosis infection. We discuss the duality between neutrophils' ability to clear infection and how increasing numbers of neutrophils contribute to active TB severity and mortality. Further investigation into the potential role of neutrophils in innate immune-mediated M. tuberculosis infection resistance is warranted since it may reveal clinically important activities for prevention as well as vaccine and treatment development.

Journal article

du Bruyn E, Peton N, Esmail H, Howlett P, Coussens AK, Wilkinson RJet al., 2018, Recent progress in understanding immune activation in the pathogenesis in HIV-TB coinfection, Current Opinion in HIV and AIDS, Vol: 13, Pages: 455-461, ISSN: 1746-630X

Purpose of review Tuberculosis is the leading infectious cause of death worldwide, and HIV-1 the best recognized risk factor for active TB. This review focuses on immune complex formation; the interplay of type I and II interferon signaling; and T-cell activation in HIV–TB pathogenesis.Recent findings Circulating immune complexes and complement, and Fcγ signaling in whole blood act as early markers of TB disease in HIV-1-infected persons. HIV-1 is associated with a type I interferon response in whole blood, reducing the specificity of TB biomarkers dependent on type I and II interferon genes. Type I and type II interferons are implicated in both protection and TB disease, a protective outcome may depend on modulating these pathways. Whilst M. tuberculosis-specific CD4 T cells are preferentially depleted during HIV-1 infection, activation markers on M. tuberculosis-specific CD4 T cells, in particular HLA-DR, reflect immune activation and have promise as biomarkers of M. tuberculosis disease activity in individuals with HIV-1.Summary TB pathogenesis in HIV-1 involves a complex interaction of underlying activation of both the innate and adaptive immune systems. Further research is required to understand whether biomarkers of activation could be used to predict or quantify TB disease in the context of HIV-1 infection.

Journal article

Singhania A, Wilkinson RJ, Rodrigue M, Haldar P, O'Garra Aet al., 2018, The value of transcriptomics in advancing knowledge of the immune response and diagnosis in tuberculosis, Nature Immunology, Vol: 19, Pages: 1159-1168, ISSN: 1529-2908

Blood transcriptomics analysis of tuberculosis has revealed an interferon-inducible gene signature that diminishes in expression after successful treatment; this promises improved diagnostics and treatment monitoring, which are essential for the eradication of tuberculosis. Sensitive radiography revealing lung abnormalities and blood transcriptomics have demonstrated heterogeneity in patients with active tuberculosis and exposed asymptomatic people with latent tuberculosis, suggestive of a continuum of infection and immune states. Here we describe the immune response to infection with Mycobacterium tuberculosis revealed through the use of transcriptomics, as well as differences among clinical phenotypes of infection that might provide information on temporal changes in host immunity associated with evolving infection. We also review the diverse blood transcriptional signatures, composed of small sets of genes, that have been proposed for the diagnosis of tuberculosis and the identification of at-risk asymptomatic people and suggest novel approaches for the development of such biomarkers for clinical use.

Journal article

Walker N, Stek C, Wasserman S, Wilkinson R, Meintjes Get al., 2018, The tuberculosis-associated immune reconstitution inflammatory syndrome: recent advances in clinical and pathogenesis research, Current Opinion in HIV and AIDS, Vol: 13, Pages: 512-521, ISSN: 1746-630X

Purpose of review Antiretroviral therapy (ART) is an essential, life-saving intervention for HIV infection. However, ART initiation is frequently complicated by the tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) in TB endemic settings. Here, we summarize the current understanding highlighting the recent evidence.Recent findings The incidence of paradoxical TB-IRIS is estimated at 18% (95% CI 16–21%), higher than previously reported and may be over 50% in high-risk groups. Early ART initiation in TB patients increases TB-IRIS risk by greater than two-fold, but is critical in TB patients with CD4 counts less than 50 cells/μl because it improves survival. There remains no validated diagnostic test for TB-IRIS, and biomarkers recently proposed are not routinely used. Prednisone initiated alongside ART in selected patients with CD4 less than 100 cells/μl reduced the risk of paradoxical TB-IRIS by 30% in a recent randomized-controlled trial (RCT) and was not associated with significant adverse effects. Effective also for treating paradoxical TB-IRIS, corticosteroids remain the only therapeutic intervention for TB-IRIS supported by RCT trial data. TB-IRIS pathogenesis studies implicate high antigen burden, innate immune cell cytotoxicity, inflammasome activation and dysregulated matrix metalloproteinases in the development of the condition.Summary Specific biomarkers would aid in identifying high-risk patients for interventions and a diagnostic test is needed. Clinicians should consider prednisone for TB-IRIS prevention in selected patients. Future research should focus on improving diagnosis and investigating novel therapeutic interventions, especially for patients in whom corticosteroid therapy is contraindicated.

Journal article

Stek C, Allwood B, Walker NF, Wilkinson RJ, Lynen L, Meintjes Get al., 2018, The Immune Mechanisms of Lung Parenchymal Damage in Tuberculosis and the Role of Host-Directed Therapy, FRONTIERS IN MICROBIOLOGY, Vol: 9, ISSN: 1664-302X

Impaired lung function is common in people with a history of tuberculosis. Host-directed therapy added to tuberculosis treatment may reduce lung damage and result in improved lung function. An understanding of the pathogenesis of pulmonary damage in TB is fundamental to successfully predicting which interventions could be beneficial. In this review, we describe the different features of TB immunopathology that lead to impaired lung function, namely cavities, bronchiectasis, and fibrosis. We discuss the immunological processes that cause lung damage, focusing on studies performed in humans, and using chest radiograph abnormalities as a marker for pulmonary damage. We highlight the roles of matrix metalloproteinases, neutrophils, eicosanoids and cytokines, like tumor necrosis factor-α and interleukin 1β, as well as the role of HIV co-infection. Finally, we focus on various existing drugs that affect one or more of the immunological mediators of lung damage and could therefore play a role as host-directed therapy.

Journal article

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