Sensitive and rapid visual detection of Orientia tsutsugamushi with Recombinase assisted dipstick detection platform

Orientia tsutsugamushi, the causative agent of scrub typhus, poses a global threat, particularly in rural areas where standard PCR methods are impractical. To enable timely management of scrub typhus, we developed a rapid, simple, sensitive, and specific molecular detection method suitable for low-resource settings with minimal instrumentation. The developed platform enabled rapid detection of OT (≤25 minutes). For Gilliam and Karp strain of OT, the limit of detection was ≥1 gene copy, with 96.9 % clinical specificity and 100 % sensitivity. Primer artefact removal reduced background noise. The minimal concentration detected 91 ag µL^-1 for Karp and 3 ag µL^-1 for Gilliam genome. The assay showed no cross-reactivity with other acute febrile or acute encephalitis syndrome related etiologies. The developed assay holds considerable promise for clinicians, facilitating prompt diagnosis and management of ST cases in settings with constrained resources.