Diagnostic challenges in re-emerging rickettsioses: why current tools fall short

SUMMARYRickettsial diseases, encompassing scrub typhus, spotted fever group rickettsioses, and typhus group rickettsioses, represent a significant and escalating public health threat worldwide, particularly in the Asia-Pacific and sub-Saharan African regions. Despite their high morbidity and potential for fatal outcomes if left untreated, these infections remain notoriously underdiagnosed due to their nonspecific clinical presentation, which frequently overlaps with other acute undifferentiated febrile illnesses (AUFIs) such as dengue, malaria, and leptospirosis. This review evaluates the evolving diagnostic landscape, highlighting the severe limitations of conventional methods: the Weil-Felix test lacks necessary specificity, while the gold standard indirect immunofluorescence assay (IFA) is primarily retrospective due to delayed seroconversion. Molecular diagnostics, particularly multiplex polymerase chain reaction (mPCR), have emerged as a critical advancement, enabling early, species-specific identification during the acute phase of infection when doxycycline therapy is most effective. We further explore the paradigm shift toward syndromic molecular panels, such as the TaqMan Array Card (TAC), which facilitate simultaneous screening for multiple AUFI pathogens. Emerging platforms, including digital PCR (dPCR) for absolute quantification and CRISPR-Cas-based point-of-care (POC) systems (SHERLOCK and DETECTR), offer promising solutions for low-resource settings. Finally, this review underscores the necessity of integrating molecular surveillance within a One Health framework and utilizing artificial intelligence (AI) to address technical and implementation barriers. Overcoming these challenges is essential for transforming rickettsial diagnosis from a reactive to a proactive strategy, ultimately reducing the global burden of these neglected zoonoses.