Development of dual detection platforms based on catalytic hairpin assembly for rapid and sensitive detection of Chlamydia trachomatis 16S rRNA.

Chlamydia trachomatis (CT) infections often remain asymptomatic yet can cause severe complications. Although nucleic acid amplification tests (NAATs) offer high sensitivity, they require expensive equipment unavailable in resource-limited settings. We developed the first application of catalytic hairpin assembly (CHA) technology combined with fluorescence immunochromatography assay (FICA) and colloidal gold immunochromatography assay (GICA) for CT 16S rRNA detection. We optimized reaction conditions (temperature, pH, probe ratio, and concentration) to minimize background signals and ensure CHA reaction feasibility. Using RT-qPCR as reference standard, we evaluated 38 CT-positive and 62 CT-negative clinical vaginal swab specimens. CHA-FICA achieved a detection limit of 10 fM with 89.47 % sensitivity and 100 % specificity within 25 min. CHA-GICA reached a detection limit of 1 pM with 81.57 % sensitivity and 100 % specificity within 30 min. Both platforms showed excellent concordance with RT-qPCR results: CHA-FICA achieved 96 % accuracy (AUC = 0.988) and CHA-GICA 93 % accuracy (AUC = 0.979). The probes remained stable for up to 5 weeks at -20 °C. This cost-effective nucleic acid detection strategy is suitable for resource-limited settings, providing both theoretical insights and practical tools for early CT diagnosis.