Yanli Zhang ¹ Xuhan Chen ² Yeling Zhong ³ Guo Fei ¹ Guifang Ouyang ^1* Rui Mao ^2,4*

• ¹ Ningbo First Hospital, Ningbo, Zhejiang Province, China
• ² Ningbo Institute of Life and Health Industry, University of Chinese Academy of Sciences, Ningbo, Zhejiang Province, China
• ³ Zhenhai People's Hospital, Ningbo, Zhejiang Province, China
• ⁴ Department of Experimental Medical Science, Ningbo No. 2 Hospital, Ningbo, China

Candida albicans, a human fungal pathogen, multiplies to invade body cells and causes fungal diseases in the condition of insufficient body's immune function. Early detection of C. albicans is required to guide appropriate prevention and treatment. The purpose of this study was to establish a C. albicans assay based on newly developed closed dumbbell-mediated isothermal amplification (CDA) to achieve rapid and simple point of care diagnostic. The CDA technique was carried out by specific primers targeting at the conserved C. albicans ITS2 gene. All primers were Tu et al., 2024). However, the drawbacks of LAMP lie in that sophisticated, cumbersome, and complex primer design, which make further expansion and application inconvenient (Gonç alves Dda et al., 2014; Mao et al., 2018;Gomez-Gutierrez and Goodwin, 2022). Thus, a novel isothermal nucleic acid amplification method with simpler primer design and more cost-effective for rapid and sensitive detection of C. albicans is needed.Closed dumbbell-mediated isothermal amplification (CDA), based on self-primer mediated single nucleic acid annealing and extending which was characterized by efficiency, high sensitivity, and high specificity (Gui et al., 2022; Mao et al., 2022;Zhang et al., 2024). In this study, the conserved internal transcribed spacer 2 (ITS2), a region between 5.8S and 28S ribosomal DNA of C. albicans was selected as the target for specific identification and amplification. The specificity and sensitivity of the developed CDA were compared with those of real-time quantitative PCR (qPCR) under similar situation. In addition, we also successfully assessed the practical application of CDA assay in real-time fluorescence and on-site visible detection of C. albicans with 100% sensitivity and specificity. All results showed the established CDA assay for C. albicans identification had the potential to achieve efficient, rapid, accurate, and visible on-site detection.