Dual one-step recombinase-aided PCR for rapid detection of Candida in blood

• DOI: 10.1007/s00253-025-13452-y
• Publication Date: 2025-03-23T02:43:02Z
• Authors: Xiaona Lyu; Kenan Peng; Zhiqiang Han; Hongyi Li; Xiaoping Chen; Shijue Gao; Yanqing Tie; Yuan Gao; Yuxin Wang; Jie Wang; Xinxin Shen; Xuejun Ma; Zhishan Feng

This study presents a novel dual one-step recombinase-aided PCR (DO-RAP) method, combined with recombinant human mannan-binding lectin protein (rhMBL; i.e., M1 protein)-conjugated magnetic bead (M1 bead) enrichment, for the early detection of Candida krusei and parapsilosis bloodstream infections. Unlike previous studies that utilized characteristic docosane being solid at room temperature melting above 44 °C as an impermeable barrier to separate two reaction steps, DO-RAP simplifies process by eliminating this step. Specificity tests genomic DNAs from 11 bacterial strains 3 fungi related infections (BSIs) confirmed no cross-reactivity, while sensitivity analysis demonstrated limits 1 copy/μL plasmids containing 26S ribosomal RNA gene fragment C. NADH5 mitochondrial 10⁻⁷ ng/μL standard parapsilosis. In simulated infection samples enriched beads, achieved thresholds CFU/mL (colony-forming unit per milliliter) in within 3.5 h, surpassing quantitative (qPCR) performance, which has 3–5 CFU/mL. Clinical validation showed strong agreement between qPCR, Kappa values 0.936 0.904 (P < 0.05). integrated approach improves speed sensitivity, eliminates need culturing, offers more efficient alternative qPCR diagnosing invasive • The method achieves CFU/mL, conventional qPCR. docosane, streamlining operations, accelerating detection. enrichment enhances pathogen capture, facilitating rapid diagnosis.