Optimization of the Simple One-Step Stool Processing Method to Diagnose Tuberculosis: Evaluation of Robustness and Stool Transport Conditions for Global Implementation.

Stool is recommended as an alternative specimen for the diagnosis of tuberculosis (TB) in young children, as they cannot easily produce sputum. The Simple One-Step (SOS) stool processing method is a new and simple stool processing method for the detection of Mycobacterium tuberculosis (MTB) using Xpert MTB/RIF Ultra (Xpert-Ultra). We determined the robustness of the SOS stool processing method and stool specimen transport conditions in participants with confirmed TB. We processed stool using the standard protocol after simulated "transport," varying time, and temperature, and experimented with slightly modified processing steps. We included 2,963 Xpert-Ultra test results from 132 stool specimens of 47 TB participants, including 11 children aged <10 years. We compared Xpert-Ultra processing errors and MTB positivity rates between standard and modified procedures. Minor deviations from the standard SOS protocol did not significantly impact the Xpert-Ultra test outcomes. The rate of Xpert-Ultra processing errors significantly increased with noncold-chain transport, exposure of stool to sample reagent at room temperature or beyond 12 h, and adding >0.8 g of stool. We found that almost all steps in the current SOS stool processing method provide optimal Xpert-Ultra results but recommend an adjustment to use a wider range of stool amounts (0.3 to 0.8 g) than advised previously (0.8 g). With this adaptation, stool-based diagnosis of TB using the SOS stool processing method can be scaled-up. The manuscript will support the global implementation and scale-up of the SOS stool method in routine settings. It also provides important insights on the optimal stool transport conditions and robustness of the SOS method, which can be used for bacteriological diagnosis of TB in children at the lowest levels of the healthcare system, avoiding lengthy healthcare-seeking pathways and additional costs.