Authors
Yuanzhao Hu, Xianyao Zhang, Changhua He, Hongliang Wang, Haimei Mao, Yi Wan, Zhiqing Yang, Yingzi Lin
Published in
The Analyst. Sep 14, 2026. Epub Sep 14, 2026.
Abstract
Methicillin-resistant Staphylococcus aureus (MRSA) is commonly identified by detection of the mecA resistance gene, but reliance on a single marker can lead to false positives, since mecA also occurs in non-MRSA organisms. Here, we report an i̲socaudomer-assisted A̲N̲D̲-gate C̲a̲s̲12a biosensor (iCasAND) for improved discrimination of MRSA by detecting the S. aureus-specific nuc gene and the methicillin-resistance gene mecA. Amplicons of both genes are digested by isocaudomer restriction enzymes (BglII and BamHI) to generate complementary sticky ends that self-assemble into split dsDNA activators of Cas12a trans-cleavage. Fluorescence is produced only when both genes are present, satisfying the AND logic condition, while T4 DNA ligase promotes complete activator formation, enhancing the signal-to-background ratio. The iCasAND system achieves a detection limit of 7682.45 ag μL-1 and a sensitivity of 45.28 CFU mL-1, discriminates MRSA from closely related pathogens, and demonstrates 100% concordance in samples from an artificial tilapia infection model. This enzyme-assisted, logic-controlled Cas12a strategy enhances specificity without requiring multiplexed crRNAs and simplifies reagents and workflow, offering a highly specific, programmable, multi-step biosensing strategy for the detection of multidrug- resistant bacteria.
PMID:
42734630
Bibliographic data and abstract were imported from PubMed on 14 Sep 2026.
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