Development of a four-gene host signature for paucibacillary TB among symptomatic individuals with sputum Xpert MTB/RIF Ultra very low and trace results.

OBJECTIVES

The Xpert MTB/RIF Ultra (Ultra) assay is widely used to diagnose pulmonary tuberculosis (TB), but "very low" or "trace" results may reflect either paucibacillary TB or TB-negative disease, complicating clinical decision-making. We evaluated host transcriptomic signatures to identify culture-confirmed paucibacillary TB among individuals with Ultra very low or trace results.

METHODS

We performed whole blood targeted transcriptional profiling of 90 symptomatic adults from Uganda, Kenya, and South Africa with Ultra very low/trace sputum results. An 81-gene customized NanoString panel representing 13 published TB signatures was analyzed using machine learning to derive a novel four-gene host signature ("TRACE4") predicting MGIT and LJ culture positivity. Validation included individuals with other respiratory diseases (n = 18) and North American-TB-negative controls (n = 20). Data were randomly split 75/25 into training (n = 67) and test (n = 23) sets. Diagnostic performance was evaluated against WHO targets.

RESULTS

TRACE4 outperformed all published signatures in both training (AUC 0.89) and test sets (AUC 0.88), achieving 82% specificity at 75% sensitivity. It also exceeded reclassification based on prior TB history (specificity 0.58 (95% CI: 0.45-0.69); sensitivity 0.35 (95% CI: 0.15-0.61)). TRACE4 showed 100% specificity in non-TB controls.

CONCLUSION

TRACE4 shows promise for identifying paucibacillary culture-positive pulmonary TB in this diagnostically challenging group.