A model based on the combination of ifn-γ, ip-10, ferritin and 25-hydroxyvitamin d for discriminating latent from active tuberculosis in children

Altres ajuts: This work was supported by grants from: (i) the Isolana Foundation, (ii) the Maria Francisca de Roviralta Foundation expedient AT/MA 1-19/07/2017, AT/MA 70-27/04/2016, and AT/MA 3-22/10/2014 and (vi) Fundació Recerca i Docència Mútua Terrassa.In recent years, pediatric research on tuberculosis (TB) has focused on addressing new biomarkers with the potential to be used as immunological non-sputum-based methods for the diagnosis of TB in children. The aim of this study was to characterize a set of cytokines and a series of individual factors (ferritin, 25-hydroxyvitamin D [25(OH)D], parasite infections, and nutritional status) to assess different patterns for discriminating between active TB and latent TB infection (LTBI) in children. The levels of 13 cytokines in QuantiFERON-TB Gold In-Tube (QFT-GIT) supernatants were analyzed in 166 children: 74 with active TB, 37 with LTBI, and 55 uninfected controls. All cytokines were quantified using Luminex or ELISA. Ferritin and 25(OH)D were also evaluated using CLIA, and Toxocara canis Ig-G antibodies were detected with a commercial ELISA kit. The combination of IP-10, IFN-γ, ferritin, and 25(OH)D achieved the best diagnostic performance to discriminate between active TB and LTBI cases in children in relation to the area under receiver operating characteristic (ROC) curve 0.955 (confidence interval 95%: 0.91-1.00), achieving optimal sensitivity and specificity for the development of a new test (93.2 and 90.0%, respectively). Children with TB showed higher ferritin levels and an inverse correlation between 25(OH)D and IFN-γ levels. The model proposed includes a combination of biomarkers for discriminating between active TB and LTBI in children to improve the accuracy of TB diagnosis in children. This combination of biomarkers might have potential for identifying the onset of primary TB in children

Urine NMR-based TB metabolic fingerprinting for the diagnosis of TB in children

Tuberculosis (TB) is a major cause of morbidity and mortality in children, and early diagnosis and treatment are crucial to reduce long-term morbidity and mortality. In this study, we explore whether urine nuclear magnetic resonance (NMR)-based metabolomics could be used to identify differences in the metabolic response of children with different diagnostic certainty of TB. We included 62 children with signs and symptoms of TB and 55 apparently healthy children. Six of the children with presumptive TB had bacteriologically confirmed TB, 52 children with unconfirmed TB, and 4 children with unlikely TB. Urine metabolic fingerprints were identified using high- and low-field proton NMR platforms and assessed with pattern recognition techniques such as principal components analysis and partial least squares discriminant analysis. We observed differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (p = 0.041 and p = 0.013, respectively). Moreover, children with unconfirmed TB with X-rays compatible with TB showed differences in the metabolic fingerprint compared to children with non-pathological X-rays (p = 0.009). Differences in the metabolic fingerprint in children with different diagnostic certainty of TB could contribute to a more accurate characterisation of TB in the paediatric population. The use of metabolomics could be useful to improve the prediction of TB progression and diagnosis in children

Perspectives for systems biology in the management of tuberculosis

Standardised management of tuberculosis may soon be replaced by individualised, precision medicine-guided therapies informed with knowledge provided by the field of systems biology. Systems biology is a rapidly expanding field of computational and mathematical analysis and modelling of complex biological systems that can provide insights into mechanisms underlying tuberculosis, identify novel biomarkers, and help to optimise prevention, diagnosis and treatment of disease. These advances are critically important in the context of the evolving epidemic of drug-resistant tuberculosis. Here, we review the available evidence on the role of systems biology approaches - human and mycobacterial genomics and transcriptomics, proteomics, lipidomics/metabolomics, immunophenotyping, systems pharmacology and gut microbiomes - in the management of tuberculosis including prediction of risk for disease progression, severity of mycobacterial virulence and drug resistance, adverse events, comorbidities, response to therapy and treatment outcomes. Application of the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach demonstrated that at present most of the studies provide "very low" certainty of evidence for answering clinically relevant questions. Further studies in large prospective cohorts of patients, including randomised clinical trials, are necessary to assess the applicability of the findings in tuberculosis prevention and more efficient clinical management of patients.Publisher PDFPeer reviewe

Diagnostic Performance of the Fujifilm SILVAMP TB-LAM in Children with Presumptive Tuberculosis

Current diagnostics for tuberculosis (TB) only manage to confirm a small proportion of children with TB and require respiratory samples, which are difficult to obtain. There is a need for non-invasive biomarker-based tests as an alternative to sputum testing. Fujifilm SILVAMP TB lipoarabinomannan (FujiLAM), a lateral-flow test to detect lipoarabinomannan in urine, is a novel non-sputum-based point-of-care diagnostic reported to have increased sensitivity for the diagnosis of TB among human immunodeficiency virus (HIV)-infected adults. We evaluate the performance of FujiLAM in children with presumptive TB. Fifty-nine children attending a paediatric hospital in Haiti with compatible signs and symptoms of TB were examined using Xpert MTB/RIF, smear microscopy and X-rays, and classified according to the certainty of diagnosis into bacteriologically confirmed TB ( = 5), unconfirmed TB (bacteriologically negative, = 50) and unlikely TB ( = 4). Healthy children ( = 20) were enrolled as controls. FujiLAM sensitivity and specificity were 60% and 95% among children with confirmed TB. FujiLAM's high specificity and its characteristics as a point-of-care indicate the test has a good potential for the diagnosis of TB in children

Urine NMR-based TB metabolic fingerprinting for the diagnosis of TB in children

Tuberculosis (TB) is a major cause of morbidity and mortality in children, and early diagnosis and treatment are crucial to reduce long-term morbidity and mortality. In this study, we explore whether urine nuclear magnetic resonance (NMR)-based metabolomics could be used to identify differences in the metabolic response of children with different diagnostic certainty of TB. We included 62 children with signs and symptoms of TB and 55 apparently healthy children. Six of the children with presumptive TB had bacteriologically confirmed TB, 52 children with unconfirmed TB, and 4 children with unlikely TB. Urine metabolic fingerprints were identified using high- and low-field proton NMR platforms and assessed with pattern recognition techniques such as principal components analysis and partial least squares discriminant analysis. We observed differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (p = 0.041 and p = 0.013, respectively). Moreover, children with unconfirmed TB with X-rays compatible with TB showed differences in the metabolic fingerprint compared to children with non-pathological X-rays (p = 0.009). Differences in the metabolic fingerprint in children with different diagnostic certainty of TB could contribute to a more accurate characterisation of TB in the paediatric population. The use of metabolomics could be useful to improve the prediction of TB progression and diagnosis in children

Discovery and validation of an NMR-based metabolomic profile in urine as TB biomarker

Despite efforts to improve tuberculosis (TB) detection, limitations in access, quality and timeliness of diagnostic services in low- and middle-income countries are challenging for current TB diagnostics. This study aimed to identify and characterise a metabolic profile of TB in urine by high-field nuclear magnetic resonance (NMR) spectrometry and assess whether the TB metabolic profile is also detected by a low-field benchtop NMR spectrometer. We included 189 patients with tuberculosis, 42 patients with pneumococcal pneumonia, 61 individuals infected with latent tuberculosis and 40 uninfected individuals. We acquired the urine spectra from high and low-field NMR. We characterised a TB metabolic fingerprint from the Principal Component Analysis. We developed a classification model from the Partial Least Squares-Discriminant Analysis and evaluated its performance. We identified a metabolic fingerprint of 31 chemical shift regions assigned to eight metabolites (aminoadipic acid, citrate, creatine, creatinine, glucose, mannitol, phenylalanine, and hippurate). The model developed using low-field NMR urine spectra correctly classified 87.32%, 85.21% and 100% of the TB patients compared to pneumococcal pneumonia patients, LTBI and uninfected individuals, respectively. The model validation correctly classified 84.10% of the TB patients. We have identified and characterised a metabolic profile of TB in urine from a high-field NMR spectrometer and have also detected it using a low-field benchtop NMR spectrometer. The models developed from the metabolic profile of TB identified by both NMR technologies were able to discriminate TB patients from the rest of the study groups and the results were not influenced by anti-TB treatment or TB location. This provides a new approach in the search for possible biomarkers for the diagnosis of TB

Urine NMR-based TB metabolic fingerprinting for the diagnosis of TB in children

Tuberculosis (TB) is a major cause of morbidity and mortality in children, and early diagnosis and treatment are crucial to reduce long-term morbidity and mortality. In this study, we explore whether urine nuclear magnetic resonance (NMR)-based metabolomics could be used to identify differences in the metabolic response of children with different diagnostic certainty of TB. We included 62 children with signs and symptoms of TB and 55 apparently healthy children. Six of the children with presumptive TB had bacteriologically confirmed TB, 52 children with unconfirmed TB, and 4 children with unlikely TB. Urine metabolic fingerprints were identified using high- and low-field proton NMR platforms and assessed with pattern recognition techniques such as principal components analysis and partial least squares discriminant analysis. We observed differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (p = 0.041 and p = 0.013, respectively). Moreover, children with unconfirmed TB with X-rays compatible with TB showed differences in the metabolic fingerprint compared to children with non-pathological X-rays (p = 0.009). Differences in the metabolic fingerprint in children with different diagnostic certainty of TB could contribute to a more accurate characterisation of TB in the paediatric population. The use of metabolomics could be useful to improve the prediction of TB progression and diagnosis in children

New approaches for the future of tuberculosis diagnosis. Interpreting conversations between the host and Mycobacterium tuberculosis

La tuberculosis (TB) es una enfermedad de transmisión aérea causada Mycobacterium tuberculosis. Aunque la TB es una enfermedad curable, es la segunda causa de muerte mundial por un agente infeccioso por detrás de la COVID-19. En 2020, alrededor del 41,4% de los pacientes con TB no se diagnosticaron ni se notificaron, contribuyendo a los 1,5 millones de muertes mundiales causadas por TB. La irrupción de la COVID-19 ha revertido años de progreso mundial aumentando la mortalidad causada por TB (anexo 1). Es necesario desarrollar métodos de diagnóstico de TB precisos, alternativos al esputo y accesibles especialmente en los países de ingresos bajos y medios, donde reside la mayor carga de la enfermedad. Actualmente no existe una prueba de referencia de TB pediátrica con suficiente sensibilidad diagnóstica (anexo 2). El artículo 1 describe un conjunto de biomarcadores identificados en suero y plasma mediante inmunoensayos para evaluar diferentes patrones discriminantes entre la infección y la enfermedad por M. tuberculosis en niños (anexo 5). La combinación de IP-10, IFN-γ, ferritina y 25(OH)D mostró una sensibilidad y especificidad óptimas para el desarrollo de un ensayo para el diagnóstico temprano de la TB en niños (93,2 y 90,0%, respectivamente). La orina es una muestra fácilmente disponible y no invasiva, que sería especialmente útil en pacientes incapaces de expectorar y en individuos con enfermedad diseminada, extrapulmonar o no cavitaria. El artículo 2 evalúa el rendimiento del ensayo Fujifilm SILVAMP TB LAM (FujiLAM), una nueva prueba de TB basada en la orina, en pacientes con presunta TB mostrando una buena precisión del ensayo para diagnosticar la TB en pacientes seropositivos y negativos. El artículo 3 evalúa el uso de FujiLAM en niños con presunta TB y sugiere su potencial como diagnóstico en el punto de atención para la TB pediátrica. El desarrollo de las ómicas ha generado nuevos conocimientos de los mecanismos subyacentes a la enfermedad. El artículo 4 identifica un perfil metabólico de la TB en la orina mediante espectrometría de resonancia magnética nuclear (RMN) de alto campo, desarrollando modelos capaces de discriminar a los pacientes con TB. Asimismo, el perfil metabólico de la TB se detectó mediante un espectrómetro de RMN portátil de bajo campo y discriminó a los pacientes con TB sin influir en el tipo de TB o tratamiento (anexo 4). El artículo 5 usó la tecnología para identificar las diferencias en la respuesta metabólica de los niños con presunta TB, mostrando diferencias en la huella metabólica de los niños con TB bacteriológicamente confirmada y no confirmada en comparación con los niños con TB poco probable (anexo 4). El artículo 6 describe la huella metabólica urinaria de los pacientes con TB durante el tratamiento hasta su finalización. Al final del tratamiento, los pacientes tenían una huella metabólica similar a la de los controles sanos. Dado el aumento de resistencia a los fármacos anti-TB y fracasos terapéuticos, esta prueba de concepto podría considerarse en los pacientes con TB resistente a los fármacos. El avance de la biología de sistemas en relación con la TB permitiría abordar con mayor precisión cuestiones clínicamente relevantes para el futuro, como el diagnóstico precoz de la TB, la individualización de las terapias preventivas y la predicción del resultado de un régimen de tratamiento (anexo 3). Esta tesis se centra en describir y evaluar nuevos enfoques en la búsqueda de posibles biomarcadores no basados en el esputo que contribuyan a mejorar el futuro diagnóstico de la TB y la evaluación de la respuesta al tratamiento anti-TB.Tuberculosis (TB) is an airborne disease caused by Mycobacterium tuberculosis. Although TB is a curable disease, it is the second leading cause of death globally from an infectious agent after COVID-19. In 2020, about 41.4% of TB patients were not diagnosed or reported, contributing to the estimated 1.5 million annual deaths caused by TB worldwide. In addition, the irruption of COVID-19 has reversed years of global progress in reducing the number of people who die from TB (annex 1). More accurate and accessible non- sputum-based diagnostic methods need to be developed, especially in low- and middle-income countries, where the greatest burden of disease resides. There is currently no paediatric TB reference test with sufficient diagnostic sensitivity (annex 2). The first article of this thesis describes a set of cytokines and a series of individual factors identified from serum and plasma samples by immunoassays to assess different discriminant patterns between M. tuberculosis infection and disease in children (article 1 and annex 5). As a result, the combination of IP-10, IFN-γ, ferritin and 25(OH)D showed the best diagnostic performance, achieving optimal sensitivity and specificity for the development of a promising early TB diagnostic test in children (93.2 and 90.0%, respectively). Urine is a readily available and non-invasive sample, which would be especially useful in patients unable to expectorate and in individuals with disseminated, extrapulmonary, or non-cavitary disease. Article 2 evaluates the performance of the Fujifilm SILVAMP TB LAM assay (FujiLAM), a new urine-based TB test, in patients with suspected TB, showing good diagnostic accuracy to diagnose TB in seropositive and negative patients. With regard to TB risk groups, article 3 evaluates the use of FujiLAM in children with presumptive TB and suggests its potential as a point-of-care diagnostic for paediatric TB. Currently, the development of omics, has allowed the generation of knowledge on the underlying mechanisms of the disease. Article 4 identifies a metabolic profile of TB in urine using high-field nuclear magnetic resonance (NMR) spectrometry by developing models capable of discriminating TB patients. In addition, the metabolic profile of TB was detected using a low-field portable NMR spectrometer and discriminated TB patients without influencing TB location or treatment (annex 4). This technology was also applied to identify differences in the metabolic response of children with presumptive TB, showing differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (article 5 and annex 4). This finding would assist in a more accurate characterisation of TB in the paediatric population. The last study in this thesis describes the urinary metabolic fingerprint of TB patients monitored during treatment to successful completion (article 6), showing differences in the metabolic fingerprint throughout treatment, such that, at the end of treatment, patients have a similar metabolic fingerprint to that of healthy controls. Given the emerging acquisition of resistance to anti-TB drugs and the increase in treatment failures, this proof of concept could be considered for monitoring the response to treatment of patients with drug-resistant TB. Advances in systems biology in relation to TB would address more precisely clinically relevant questions for the future, such as the early diagnosis of TB, the individualisation of preventive therapies, and the prediction of the outcome of a treatment regimen (annex 3). Overall, this thesis focuses on describing and evaluating new approaches in the search for potential non- sputum-based biomarkers contributing to improve future TB diagnosis and the assessment of response to anti-TB treatment

Diagnostic Performance of the Fujifilm SILVAMP TB-LAM in Children with Presumptive Tuberculosis

Current diagnostics for tuberculosis (TB) only manage to confirm a small proportion of children with TB and require respiratory samples, which are difficult to obtain. There is a need for non-invasive biomarker-based tests as an alternative to sputum testing. Fujifilm SILVAMP TB lipoarabinomannan (FujiLAM), a lateral-flow test to detect lipoarabinomannan in urine, is a novel non-sputum-based point-of-care diagnostic reported to have increased sensitivity for the diagnosis of TB among human immunodeficiency virus (HIV)-infected adults. We evaluate the performance of FujiLAM in children with presumptive TB. Fifty-nine children attending a paediatric hospital in Haiti with compatible signs and symptoms of TB were examined using Xpert MTB/RIF, smear microscopy and X-rays, and classified according to the certainty of diagnosis into bacteriologically confirmed TB (n = 5), unconfirmed TB (bacteriologically negative, n = 50) and unlikely TB (n = 4). Healthy children (n = 20) were enrolled as controls. FujiLAM sensitivity and specificity were 60% and 95% among children with confirmed TB. FujiLAM’s high specificity and its characteristics as a point-of-care indicate the test has a good potential for the diagnosis of TB in children