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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.

Interferon-gamma (IFN-γ) release assays (IGRAs) are increasingly used to test for latent TB infection. Although highly specific, IGRAs have a relatively high false-negative rate in active TB patients. A more sensitive assay is needed. IP-10 is an alternative biomarker with a 100 fold high expression level compared to IFN-γ, allowing for different analysis platforms including molecular detection. The PCR technique is already an integrated tool in most TB laboratories and thus an obvious platform to turn to. In this case-control study, we investigated the diagnostic sensitivity and specificity of a molecular assay detecting IP-10 mRNA expression following antigen stimulation of a blood sample. We included 89 TB patients and 99 healthy controls. Blood was drawn in QuantiFERON-TB Gold In-Tube (QFT) tubes. Eight hours post-stimulation IP-10 mRNA expression was analyzed and 20 hours post-stimulation IP-10 and IFN-γ protein plasma levels were analyzed using an in-house IP-10 ELISA and the official QFT ELISA, respectively. The IP-10 mRNA assay provided a high specificity (98%) and sensitivity (80%) and AUC=0.97, however, the QFT assay provided a higher overall diagnostic potential with specificity (100%) and sensitivity (90%) and AUC=0.99. The IP-10 protein assay performed on par with the QFT assay with specificity (98%) and sensitivity (87%) and AUC=0.98. We have provided proof of high technical performance of a molecular assay detecting IP-10 mRNA expression. As a diagnostic tool, this assay would gain from further optimization especially on the kinetics of IP-10 mRNA expression.

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