Jia Xinbei, Zhang Yiqin, Zhou Lin, Zhou Juan, Xiao Fei, Ma Lijuan, Wang Yi, Tai Jun
Department of Otorhinolaryngology Head and Neck Surgery, Children's Hospital Capital Institute of Pediatrics, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100020, China.
Clinical Laboratory Center, Children's Hospital Capital Institute of Pediatrics, Beijing 100020, China.
ACS Omega. 2025 Feb 28;10(9):9768-9777. doi: 10.1021/acsomega.5c00479. eCollection 2025 Mar 11.
(group A streptococcus, GAS) is the leading bacterial cause of acute pharyngitis in children and adolescents. Rapid and reliable diagnosis of GAS pharyngitis is essential for guiding a timely antibiotic treatment. Here, we developed a rapid, highly sensitive, and specific test platform for the detection of GAS, designated GAS-MCDA-CRISPR. In this diagnostic platform, the multiple cross displacement amplification (MCDA) technique was utilized to preamplify the specific gene of GAS. Subsequently, the CRISPR-Cas12a-based biosensing system was employed to decode the MCDA products. MCDA primers, a guide RNA (gRNA), and a quenched fluorescent single-stranded DNA (ssDNA) reporter were designed to target the gene of GAS. The GAS-MCDA-CRISPR assay demonstrated the ability to detect GAS genomic DNA at a concentration as low as 45 fg per microliter while exhibiting no cross-reactivity with other non-GAS pathogens. Moreover, 56 clinical samples were correctly detected by the GAS-MCDA-CRISPR assay. These data highlighted that the GAS-MCDA-CRISPR assay is a reliable diagnostic tool for the reliable and quick diagnosis of GAS infection.
A组链球菌（GAS）是儿童和青少年急性咽炎的主要细菌性病因。GAS咽炎的快速可靠诊断对于指导及时的抗生素治疗至关重要。在此，我们开发了一种用于检测GAS的快速、高度灵敏且特异的检测平台，命名为GAS-MCDA-CRISPR。在这个诊断平台中，多重交叉置换扩增（MCDA）技术被用于预扩增GAS的特定基因。随后，基于CRISPR-Cas12a的生物传感系统被用于解码MCDA产物。MCDA引物、向导RNA（gRNA）和淬灭荧光单链DNA（ssDNA）报告分子被设计用于靶向GAS基因。GAS-MCDA-CRISPR检测方法显示出能够检测低至每微升45飞克浓度的GAS基因组DNA，同时对其他非GAS病原体无交叉反应性。此外，GAS-MCDA-CRISPR检测方法正确检测了56份临床样本。这些数据突出表明，GAS-MCDA-CRISPR检测方法是用于可靠快速诊断GAS感染的可靠诊断工具。
