Stool-Based Molecular Tuberculosis Treatment Monitoring: A Faster Means for Detecting Persistent Mycobacteria Compared to Phenotypic Culture

Abstract

Background: Tuberculosis (TB) treatment monitoring is hindered by the lack of a rapidly measured biomarker that accurately predicts clinically relevant outcomes. Symptom screening poorly correlates with bacillary burden. Although culture is a direct measure of viable bacillary burden, the long turnaround time makes it clinically irrelevant.

Methods: The TB treatment monitoring potential of stool-based, quantitative polymerase chain reaction (qPCR) was prospectively assessed among 231 participants of all ages from Eswatini, Tanzania, and Mozambique with microbiologically confirmed TB. Stool qPCR results were compared to sputum culture, persistent symptoms, drug resistance, and World Health Organization TB outcomes.

Results: Quantitative bacillary burden measured by stool qPCR strongly correlated with sputum culture at baseline (Spearman correlation r _s = 0.79; P < .001). Stool was successfully collected at >90% of all timepoints, while sputum collection decreased to <50% at the end of therapy. Participants with isoniazid or rifampin resistance demonstrated decreased bacillary clearance by sputum culture and stool qPCR during the first 2 weeks of treatment. Participants who remained culture positive at 2 months had a slower decrease in bacillary burden measured by stool qPCR compared to those who were culture negative by 2 months. The odds of a participant being culture positive at 2 months was associated with a lower initial qPCR cycle threshold (odds ratio [OR], 0.792; P = .004), and a smaller absolute difference between the qPCR cycle threshold measured at 2 weeks and baseline (OR, 0.72; P = .0006). Neither sputum culture, sputum Xpert Ultra, or stool qPCR was associated with resolution of symptoms or in-treatment death.

Conclusions: Stool-based TB treatment monitoring correlates with sputum culture but provides results faster, leverages a more accessible specimen, and identifies patients with TB who are at risk for drug resistance and persistent 2-month culture positivity. None of the quantitative tests of bacillary burden singularly could predict symptom resolution or death.

Keywords: sputum culture; stool qPCR test; stool-based; stool-based molecular test; treatment monitoring.

Graphical Abstract

Figure 1.

Overlap of microbiologic testing. Venn diagram demonstrating the overlap in positivity rate among the stool quantitative polymerase chain reaction (qPCR), sputum culture, and sputum Xpert Ultra (Xpert).

Figure 2.

Flowchart of performed and positive sputum culture, sputum Xpert Ultra (Xpert), and stool quantitative polymerase chain reaction (qPCR) assays at all study timepoints. In-treatment sputum collection stopped after 2 consecutive negative sputum Xpert tests were obtained, unless symptoms persisted. End-of-therapy sputum was collected from all participants to support assignment of World Health Organization–defined treatment outcomes. Stool qPCR testing was performed until 2 consecutive negative results were obtained.

Figure 3.

Quantitative decrease in bacillary burden with antituberculosis treatment (ATT). A, Percentage of sample collected after initiation of ATT. B–D, Quantitative bacillary burden detected by sputum culture (B), sputum Xpert Ultra (Xpert) (C), and stool quantitative polymerase chain reaction (qPCR) (D). A Wilcoxon pair-matched test evaluated the quantitative burden at baseline compared to 2 weeks for those positive at baseline and with a result reported at 2 weeks. Negative results at 2 weeks are considered to have a time to positivity (TTP) of 42 and cycle threshold (Ct) of 38.

Figure 4.

Lack of change in quantifiable decrease in bacillary burden among patients with tuberculosis who were culture positive at 2 months. Tuberculosis bacillary burden was quantified using sputum culture (A), sputum Xpert Ultra (B), and stool quantitative polymerase chain reaction (C). The quantitative changes between baseline and 2 weeks were evaluated using Mann-Whitney test. Abbreviations: ATT, anti-tuberculosis treatment; Ct, cycle threshold; qPCR, quantitative polymerase chain reaction; TTP, time to positivity.