Background 4 million tuberculosis (TB) cases are undiagnosed annually. A global host biosignature to distinguish TB from other respiratory diseases (ORD) is required to develop a point-of-care (POC) triage test for TB. Our aim was to identify a global host biosignature for TB in sputum samples.

Methods Sputum samples from HIV negative confirmed TB patients (n=221) or patients with other respiratory disease (ORD) (n=414) were analysed from South Africa, Peru, Vietnam and The Gambia. Cryopreserved native sputum samples from all countries were digested with sputolysin for 15 minutes at room temperature, centrifuged, and supernatants stored at -80oC until analysis. Multiplex cytokine arrays were used to analyse 64 host markers. Analyte levels from sputum digested after freezing were compared with those from paired freshly digested supernatants (n=45) in the Gambian cohort.

Results Levels of TNF-α (p<0.0001), MMP-2 (p<0.0001), IL-1β (p=0.0002), IL-22 (p=0.0004) and LIGHT/TNFSF14 (p=0.0007) were all significantly higher in sputum from TB compared to ORD patients. A global signature consisting of 12 analytes resulted in an AUC of 0.93, Sensitivity of 81% and Specificity of 91% while a reduced performance was seen for a 7-marker signature (AUC 0.81, Sensitivity 73% and Specificity 76%). A 4-marker signature consisting of GCSF, IL19, IL2 and MMP1 from South Africa and Gambia combined resulted in an AUC of 0.84, Sensitivity of 82% and Specificity of 70%. Fresh samples had significantly higher levels of analytes compared to post-hoc digested sputum.

Conclusion We identified a global signature that reached WHO TPP for a triage test but could not reduce from 12 markers with all countries combined. A 4-marker signature from Gambia and South Africa performed well but did not reach the TPP for a Triage test. Freshly digested sputum was superior to post-hoc digestion, indicating that use of frozen samples was a limitation in this study.