Mass Spectrometric Identification of Urinary Biomarkers of Pulmonary Tuberculosis

Abstract

BackgroundTuberculosis (TB) is the leading infectious cause of death worldwide. A major barrier to control of the pandemic is a lack of clinical biomarkers with the ability to distinguish active TB from healthy and sick controls and potential for development into point-of-care diagnostics.MethodsWe conducted a prospective case control study to identify candidate urine-based diagnostic biomarkers of active pulmonary TB (discovery cohort) and obtained a separate blinded “validation” cohort of confirmed cases of active pulmonary TB and controls with non-tuberculous pulmonary disease for validation. Clean-catch urine samples were collected and analyzed using high performance liquid chromatography-coupled time-of-flight mass spectrometry.ResultsWe discovered ten molecules from the discovery cohort with receiver-operator characteristic (ROC) area-under-the-curve (AUC) values >85%. These 10 molecules also significantly decreased after 60 days of treatment in a subset of 20 participants followed over time. Of these, a specific combination of diacetylspermine, neopterin, sialic acid, and N-acetylhexosamine exhibited ROC AUCs >80% in a blinded validation cohort of participants with active TB and non-tuberculous pulmonary disease.ConclusionUrinary levels of diacetylspermine, neopterin, sialic acid, and N-acetylhexosamine distinguished patients with tuberculosis from healthy controls and patients with non-tuberculous pulmonary diseases, providing a potential noninvasive biosignature of active TB.FundingThis study was funded by Weill Cornell Medicine, the National Institute of Allergy and Infectious Diseases, the Clinical and Translational Science Center at Weill Cornell, the NIH Fogarty International Center grants, and the NIH Tuberculosis Research Unit (Tri-I TBRU).

Urinary biomarkers of mycobacterial load and treatment response in pulmonary tuberculosis

“…Previous work from our and other groups showed elevated levels of urinary DiAcSpm in ATB cases (22,43). Additionally, our study demonstrates that DiAcSpm levels decrease rapidly with effective TB treatment.…”

“…These findings, therefore, suggest that the molecular markers are not just measuring pharmacological actions of medication on host metabolism, but more importantly are reporting on disease activity itself. Previous work reported that urinary levels of these 7 molecules were elevated in ATB cases compared with cases of non-TB pulmonary disease with an overall sensitivity and specificity of more than 80% before initiation of treatment ( 22 ). Kynurenine, neopterin, and sialic acid levels have also been previously reported to be increased in various human ATB biofluids ( 28 – 33 ).…”

“…In previous work, we identified N 1 , N 12 -diacetylspermine (DiAcSpm), hydroxykynurenine, neopterin, N -acetylhexosamine, ureidopropionic acid, sialic acid, and an uncharacterized molecule with mass-to-charge ratio ( m/z ) of 241.0903 as potential urinary biomarkers of active pulmonary TB (ATB) ( Table 1 ) ( 22 ). These urinary molecule levels decreased after 60 days of anti-TB treatment in 20 participants, supporting their utility for indicating active TB disease ( 22 ).…”

Deficiency in the glycosyltransferase Gcnt1 increases susceptibility to tuberculosis through a mechanism involving neutrophils

“…Of note, previous reports showed that the urinary levels of sialic acid, the terminal glycan of sLe x structure, discriminate patients with active TB from healthy controls or patients with non-tuberculous pulmonary diseases. 35 Also, the regulation of the gene encoding CMAS, the enzyme that provides the substrate for the addition of sialic acid, specifically changed with infection with Mtb. 36 Thus, modulation of sialylated antigens seems to occur in TB, as described in several pathologies, such as other infectious diseases and cancer.…”

Overview of Brain-to-Gut Axis Exposed to Chronic CNS Bacterial Infection(s) and a Predictive Urinary Metabolic Profile of a Brain Infected by Mycobacterium tuberculosis

“…Based on the premise that the urine contains the accumulation of all end-product metabolites of the body, logic dictates that chronic bacterial infection(s) of the CNS should, in principle, result in persistent feedback on the gut via the BGA, communicated via the CSF and blood, leading to dysbiosis and an altered urinary metabolome. In research on infectious diseases, urinary profiling has received much attention, in particular regarding pulmonary tuberculosis (TB) -a disease caused by Mycobacterium tuberculosis (Mtb) -about which several studies have been conducted using urine for the detection of clinically relevant biomarkers (Banday et al, 2011;Bonkat, 2012;Das et al, 2015;Luies and Loots, 2016;Luies et al, 2017;Preez et al, 2017;Isa et al, 2018). The detection of lipoarabinomannan (LAM), for instance, a Mycobacterium-specific liposaccharide from the Mtb cell wall, is an example of the basis of a well-studied commercial ELISA assay that shows promise for its diagnostic use in urine with a reported sensitivity of 74% and specificity of 86.9% in a study performed on 148 confirmed TB patients (Tessema et al, 2001); a sensitivity of 80.3% and specificity of 99% in a study conducted on 132 confirmed TB patients (Boehme et al, 2005); and a sensitivity of 44% and specificity of 89% in a study conducted on 195 TB-positive patients in a high-HIV prevalence setting (Mutetwa et al, 2009).…”