Imperial College London

ProfessorRobertWilkinson

Faculty of MedicineDepartment of Medicine

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

344 results found

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 DB, Wilkinson RJet al., 2019, Variation in pre-therapy levels of selected Mycobacterium tuberculosis transcripts insputumand their relationship with 2-month culture conversion, Wellcome Open Research, Vol: 4, Pages: 106-106

<ns4:p><ns4:bold>Background: </ns4:bold>The abundance of transcripts arising from <ns4:italic>Mycobacterium tuberculosis </ns4:italic>(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.</ns4:p><ns4:p> <ns4:bold>Methods: </ns4:bold>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.</ns4:p><ns4:p> <ns4:bold>Results: </ns4:bold>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.</ns4:p><ns4:p> Increased <ns4:italic>icl1 </ns4:italic>and <ns4:italic>prpD</ns4:italic> and <ns4:italic>rpsN2:rpsN1</ns4:italic> in sputum relative to culture suggested cholesterol utilization and a low-zinc environment respectively. Increased <ns4:italic>hspX</ns4:italic> and decreased <ns4:italic>atpA</ns4:italic> and <ns4:italic>nuoG</ns4:italic> relative to exponential culture suggested a slowly metabolizing subpopulation of MTB. While the the <ns4:italic>hspX</ns4:italic><ns4:sup>hi</ns4:sup><ns4:italic>atpA</ns4:italic><ns4:sup>lo</ns4:sup><ns4:italic>nuoG</ns4:italic><ns4:sup>lo</ns4:sup> signal varied, we did not obser

Journal article

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

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

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

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

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

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., 2018, A systematic review on the effect of HIV infection on the pharmacokinetics of first-line tuberculosis drugs, Clinical Pharmacokinetics, 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

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

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

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

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., Drug susceptibility testing and mortality in patients treated for tuberculosis in high-burden countries: A multi-centre cohort study, Lancet Infectious Diseases, 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

Van Der Meeren O, Hatherill M, Nduba V, Wilkinson RJ, Muyoyeta M, van Brakel E, Ayles HM, Henostroza G, Thienemann F, Scriba TJ, Diacon A, Blatner GL, Demoitié M-A, Tameris M, Malahleha M, Innes JC, Hellstrom E, Martinson N, Singh T, Akite EJ, Azam AK, Bollaerts A, Ginsberg AM, Evans TG, Gillard P, Tait DRet al., 2018, Phase 2b controlled trial of M72/AS01E vaccine to prevent tuberculosis, New England Journal of Medicine, Vol: 379, Pages: 1621-1634, ISSN: 0028-4793

BackgroundA vaccine to interrupt the transmission of tuberculosis is needed.MethodsWe conducted a randomized, double-blind, placebo-controlled, phase 2b trial of the M72/AS01E tuberculosis vaccine in Kenya, South Africa, and Zambia. Human immunodeficiency virus (HIV)–negative adults 18 to 50 years of age with latent M. tuberculosis infection (by interferon-γ release assay) were randomly assigned (in a 1:1 ratio) to receive two doses of either M72/AS01E or placebo intramuscularly 1 month apart. Most participants had previously received the bacille Calmette–Guérin vaccine. We assessed the safety of M72/AS01E and its efficacy against progression to bacteriologically confirmed active pulmonary tuberculosis disease. Clinical suspicion of tuberculosis was confirmed with sputum by means of a polymerase-chain-reaction test, mycobacterial culture, or both.ResultsWe report the primary analysis (conducted after a mean of 2.3 years of follow-up) of the ongoing trial. A total of 1786 participants received M72/AS01E and 1787 received placebo, and 1623 and 1660 participants in the respective groups were included in the according-to-protocol efficacy cohort. A total of 10 participants in the M72/AS01E group met the primary case definition (bacteriologically confirmed active pulmonary tuberculosis, with confirmation before treatment), as compared with 22 participants in the placebo group (incidence, 0.3 cases vs. 0.6 cases per 100 person-years). The vaccine efficacy was 54.0% (90% confidence interval [CI], 13.9 to 75.4; 95% CI, 2.9 to 78.2; P=0.04). Results for the total vaccinated efficacy cohort were similar (vaccine efficacy, 57.0%; 90% CI, 19.9 to 76.9; 95% CI, 9.7 to 79.5; P=0.03). There were more unsolicited reports of adverse events in the M72/AS01E group (67.4%) than in the placebo group (45.4%) within 30 days after injection, with the difference attributed mainly to injection-site reactions and influenza-like symptoms. Serious adverse events, potent

Journal article

Kendall EA, Azman AS, Maartens G, Boulle A, Wilkinson RJ, Dowdy DW, Rangaka MXet al., 2018, Projected population-wide impact of antiretroviral therapy-linked isoniazid preventive therapy in a high-burden setting, AIDS, 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

Lesosky M, Rangaka MX, Pienaar C, Coussens AK, Goliath RT, Mathee S, Mwansa-Kambafwile J, Maartens G, Wilkinson RJ, Wilkinson KAet al., 2018, Plasma biomarkers to detect prevalent, or predict progressive, HIV-1-associated tuberculosis, Clinical Infectious Diseases, 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

Sallin MA, Kauffman KD, Riou CR, Du Bruyn E, Foreman TW, Sakai S, Hoft SG, Myers TG, Gardina PJ, Sher AF, Moore R, Wilder-Kofie T, Moore IN, Sette A, Lindestam Arlehamn CS, Wilkinson RJ, Barber DLet al., 2018, Host resistance to pulmonary Mycobacterium tuberculosis 2 infection requires CD153 expression, Nature Microbiology, Vol: 3, Pages: 1198-1205, ISSN: 2058-5276

Mycobacterium tuberculosis infection (Mtb) is the leading cause of death due to a single infectious agent and is among the top ten causes of all human deaths worldwide1. CD4 T cells are essential for resistance to Mtb infection, and for decades it has been thought that IFNγ production is the primary mechanism of CD4 T-cell-mediated protection2,3. However, IFNγ responses do not correlate with host protection, and several reports demonstrate that additional anti-tuberculosis CD4 T-cell effector functions remain unaccounted for4,5,6,7,8. Here we show that the tumour-necrosis factor (TNF) superfamily molecule CD153 (encoded by the gene Tnfsf8) is required for control of pulmonary Mtb infection by CD4 T cells. In Mtb-infected mice, CD153 expression is highest on Mtb-specific T helper 1 (TH1) cells in the lung tissue parenchyma, but its induction does not require TH1 cell polarization. CD153-deficient mice develop high pulmonary bacterial loads and succumb early to Mtb infection. Reconstitution of T-cell-deficient hosts with either Tnfsf8−/− or Ifng−/− CD4 T cells alone fails to rescue mice from early mortality, but reconstitution with a mixture of Tnfsf8−/− and Ifng−/− CD4 T cells provides similar protection as wild-type T cells. In Mtb-infected non-human primates, CD153 expression is much higher on Ag-specific CD4 T cells in the airways compared to blood, and the frequency of Mtb-specific CD153-expressing CD4 T cells inversely correlates with bacterial loads in granulomas. In Mtb-infected humans, CD153 defines a subset of highly polyfunctional Mtb-specific CD4 T cells that are much more abundant in individuals with controlled latent Mtb infection compared to those with active tuberculosis. In all three species, Mtb-specific CD8 T cells did not upregulate CD153 following peptide stimulation. Thus, CD153 is a major immune mediator of host protection against pulmonary Mtb infection and CD4 T cells are one importan

Journal article

Schutz C, Davis AG, Sossen B, Lai R, Ntsekhe M, Harley YXR, Wilkinson Ret al., 2018, Corticosteroids as an adjunct to tuberculosis therapy, Expert Review of Respiratory Medicine, Vol: 12, Pages: 881-891, ISSN: 1747-6348

Introduction: Inflammation, or the prolonged resolution of inflammation, contributes to death from tuberculosis. Interest in inflammatory mechanisms and the prospect of beneficial immune modulation as an adjunct to antibacterial therapy has revived and the concept of host directed therapies has been advanced. Such renewed attention has however, overlooked the experience of such therapy with corticosteroids.Areas covered: The authors conducted literature searches and evaluated randomized clinical trials, systematic reviews and current guidelines and summarize these findings. They found evidence of benefit in meningeal and pericardial tuberculosis in HIV-1 uninfected persons, but less so in those HIV-1 coinfected and evidence of harm in the form of opportunist malignancy in those not prescribed antiretroviral therapy. Adjunctive corticosteroids are however of benefit in the treatment and prevention of paradoxical HIV-tuberculosis immune reconstitution inflammatory syndrome.Expert commentary: Further high-quality clinical trials and experimental medicine studies are warranted and analysis of materials arising from such studies could illuminate ways to improve corticosteroid efficacy or identify novel pathways for more specific intervention.

Journal article

Day CL, Abrahams DA, Bunjun R, Stone L, de Kock M, Walzl G, Wilkinson RJ, Burgers WA, Hanekom WAet al., 2018, PD-1 expression on mycobacterium tuberculosis-specific CD4 T cells is associated with bacterial load in human tuberculosis, Frontiers in Immunology, Vol: 9, ISSN: 1664-3224

Persistent antigen stimulation in chronic infections has been associated with antigen-specific T cell dysfunction and upregulation of inhibitory receptors, including programmed cell death protein 1 (PD-1). Pulmonary tuberculosis (TB) disease is characterized by high levels of Mycobacterium tuberculosis (Mtb), yet the relationship between bacterial load, PD-1 expression, and Mtb-specific T cell function in human TB has not been well-defined. Using peripheral blood samples from adults with LTBI and with pulmonary TB disease, we tested the hypothesis that PD-1 expression is associated with bacterial load and functional capacity of Mtb-specific T cell responses. We found that PD-1 was expressed at significantly higher levels on Th1 cytokine-producing Mtb-specific CD4 T cells from patients with smear-positive TB, compared with smear-negative TB and LTBI, which decreased after completion of anti-TB treatment. By contrast, expression of PD-1 on Mtb-specific CD8 T cells was significantly lower than on Mtb-specific CD4 T cells and did not differ by Mtb infection and disease status. In vitro stimulation of PBMC with Mtb antigens demonstrated that PD-1 is induced on proliferating Mtb-specific CD4 T cells and that Th1 cytokine production capacity is preferentially maintained within PD-1+ proliferating CD4 T cells, compared with proliferating Mtb-specific CD4 T cells that lack PD-1 expression. Together, these data indicate that expression of PD-1 on Mtb-specific CD4 T cells is indicative of mycobacterial antigen exposure and identifies a population of effector cells with Th1 cytokine production capacity. These studies provide novel insights into the role of the PD-1 pathway in regulating CD4 and CD8 T cell responses in Mtb infection and provide rationale for future studies to evaluate PD-1 expression on antigen-specific CD4 T cells as a potential biomarker for bacterial load and treatment response in human TB.

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, 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

Malherbe ST, Dupont P, Kant I, Ahlers P, Kriel M, Loxton AG, Chen RY, Via LE, Thienemann F, Wilkinson RJ, Barry CE, Griffith-Richards S, Ellman A, Ronacher K, Winter J, Walzl G, Warwick JMet al., 2018, A semi-automatic technique to quantify complex tuberculous lung lesions on F-18-fluorodeoxyglucose positron emission tomography/computerised tomography images, EJNMMI Research, Vol: 8, ISSN: 2191-219X

BackgroundThere is a growing interest in the use of 18F-FDG PET-CT to monitor tuberculosis (TB) treatment response. However, TB causes complex and widespread pathology, which is challenging to segment and quantify in a reproducible manner.To address this, we developed a technique to standardise uptake (Z-score), segment and quantify tuberculous lung lesions on PET and CT concurrently, in order to track changes over time. We used open source tools and created a MATLAB script. The technique was optimised on a training set of five pulmonary tuberculosis (PTB) cases after standard TB therapy and 15 control patients with lesion-free lungs.ResultsWe compared the proposed method to a fixed threshold (SUV > 1) and manual segmentation by two readers and piloted the technique successfully on scans of five control patients and five PTB cases (four cured and one failed treatment case), at diagnosis and after 1 and 6 months of treatment. There was a better correlation between the Z-score-based segmentation and manual segmentation than SUV > 1 and manual segmentation in terms of overall spatial overlap (measured in Dice similarity coefficient) and specificity (1 minus false positive volume fraction). However, SUV > 1 segmentation appeared more sensitive. Both the Z-score and SUV > 1 showed very low variability when measuring change over time. In addition, total glycolytic activity, calculated using segmentation by Z-score and lesion-to-background ratio, correlated well with traditional total glycolytic activity calculations. The technique quantified various PET and CT parameters, including the total glycolytic activity index, metabolic lesion volume, lesion volumes at different CT densities and combined PET and CT parameters. The quantified metrics showed a marked decrease in the cured cases, with changes already apparent at month one, but remained largely unchanged in the failed treatment case.ConclusionsOur technique is promising to segme

Journal article

Singhania A, Verma R, Graham CM, Lee J, Tran T, Richardson M, Lecine P, Leissner P, Berry MPR, Wilkinson RJ, Kaiser K, Rodrigue M, Woltmann G, Haldar P, O'Garra Aet al., 2018, A modular transcriptional signature identifies phenotypic heterogeneity of human tuberculosis infection., Nature Communications, Vol: 9, ISSN: 2041-1723

Whole blood transcriptional signatures distinguishing active tuberculosis patients from asymptomatic latently infected individuals exist. Consensus has not been achieved regarding the optimal reduced gene sets as diagnostic biomarkers that also achieve discrimination from other diseases. Here we show a blood transcriptional signature of active tuberculosis using RNA-Seq, confirming microarray results, that discriminates active tuberculosis from latently infected and healthy individuals, validating this signature in an independent cohort. Using an advanced modular approach, we utilise the information from the entire transcriptome, which includes overabundance of type I interferon-inducible genes and underabundance of IFNG and TBX21, to develop a signature that discriminates active tuberculosis patients from latently infected individuals or those with acute viral and bacterial infections. We suggest that methods targeting gene selection across multiple discriminant modules can improve the development of diagnostic biomarkers with improved performance. Finally, utilising the modular approach, we demonstrate dynamic heterogeneity in a longitudinal study of recent tuberculosis contacts.

Journal article

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

Contextual functional interpretation of -omics data derived from clinical samplesis a classical and difficult problem in computational systems biology. Themeasurement of thousands of data points on single samples has becomeroutine but relating ‘big data’ datasets to the complexities of humanpathobiology is an area of ongoing research. Complicating this is the fact thatmany publically available datasets use bulk transcriptomics data from complextissues 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 multiscaleanalysis), a network-based data integration method using clinical phenotypeand microarray data as inputs. ANIMA is implemented in R and Neo4j and runsin Docker containers. In short, the build algorithm iterates over one or moretranscriptomics datasets to generate a large, multipartite association networkby 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 directlyusing Cypher, or via custom functions that communicate with the graphdatabase via language-specific APIs. We developed a web application usingShiny, which provides fully interactive, multiscale views of the data. Using ourapproach, we show that we can reconstruct multiple features of disease statesat various scales of organization, from transcript abundance patterns ofindividual genes through co-expression patterns of groups of genes to patternsof cellular behaviour in whole blood samples, both in single experiments as wellas in a meta-analysis of multiple datasets.

Journal article

Barr DA, Kerkhoff AD, Schuttz C, Ward AM, Davies GR, Wilkinson RJ, Meintjes Get al., 2018, HIV-associated M. tuberculosis blood stream infection is under-diagnosed by a single blood culture, Journal of Clinical Microbiology, Vol: 56, ISSN: 0095-1137

We assessed the additional diagnostic yield for Mycobacterium tuberculosis bloodstream infection (BSI) by doing more than one tuberculosis (TB) blood culture from HIV-infected inpatients. In a retrospective analysis of two cohorts based in Cape Town, South Africa, 72/99 (73%) patients with M. tuberculosis BSI were identified by the first of two blood cultures during the same admission, with 27/99 (27%; 95% confidence interval [CI], 18 to 36%) testing negative on the first culture but positive on the second. In a prospective evaluation of up to 6 blood cultures over 24 h, 9 of 14 (65%) patients with M. tuberculosis BSI had M. tuberculosis grow on their first blood culture; 3 more patients (21%) were identified by a second independent blood culture at the same time point, and the remaining 2 were diagnosed only on the 4th and 6th blood cultures. Additional blood cultures increase the yield for M. tuberculosis BSI, similar to what is reported for nonmycobacterial BSI.

Journal article

Lowe D, Demaret J, Bangani N, Nakiwala JK, Goliath RT, Wilkinson K, Wilkinson R, Martineau ARet al., 2018, Differential effect of viable versus necrotic neutrophils on Mycobacterium tuberculosis growth and cytokine induction in whole blood, Frontiers in Immunology, Vol: 9, ISSN: 1664-3224

Neutrophils exert both positive and negative influences on the host response to tuberculosis, but the mechanisms by which these differential effects are mediated are unknown. We studied the impact of live and dead neutrophils on the control of Mycobacterium tuberculosis using a whole blood bioluminescence-based assay, and assayed supernatant cytokine concentrations using Luminex™ technology and ELISA. CD15+ granulocyte depletion from blood prior to infection with M. tuberculosis-lux impaired control of mycobacteria by 96 h, with a greater effect than depletion of CD4+, CD8+, or CD14+ cells (p < 0.001). Augmentation of blood with viable granulocytes significantly improved control of mycobacteria by 96 h (p = 0.001), but augmentation with necrotic granulocytes had the opposite effect (p = 0.01). Both augmentations decreased supernatant concentrations of tumor necrosis factor and interleukin (IL)-12 p40/p70, but necrotic granulocyte augmentation also increased concentrations of IL-10, G-CSF, GM-CSF, and CCL2. Necrotic neutrophil augmentation reduced phagocytosis of FITC-labeled M. bovis BCG by all phagocytes, whereas viable neutrophil augmentation specifically reduced early uptake by CD14+ cells. The immunosuppressive effect of dead neutrophils required necrotic debris rather than supernatant. We conclude that viable neutrophils enhance control of M. tuberculosis in blood, but necrotic neutrophils have the opposite effect—the latter associated with induction of IL-10, growth factors, and chemoattractants. Our findings suggest a mechanism by which necrotic neutrophils may exert detrimental effects on the host response in active tuberculosis.

Journal article

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