RPA-CRISPR/Cas12a detection based on HCMV-UL123 gene: A way with higher detection rate than clinical detection methods

IF 4.7 3区医学 Q1 INFECTIOUS DISEASES

Journal of Infection and Public Health Pub Date : 2025-05-28 DOI:10.1016/j.jiph.2025.102845

Tianyang Liu , Yao Wang , Zihao Liao , Xiaowen Li , Shanshan Tang , Zhongming Zhang , Ming Chu , Lanlan Wei

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引用次数: 0

Abstract

Objective

Human cytomegalovirus (HCMV), a prevalent double-stranded DNA enveloped virus, poses a threat to immunocompromised individuals. The current clinical detection methods are insufficient in sensitivity, highlighting the need for more effective approaches.

Methods

We designed and screened RPA primers and crRNA based on the UL123 gene of HCMV. Evaluate the HCMV-RPA-CRISPR detection method using cloned plasmids and the whole-genome samples of HCMV-infected cells. Conduct RPA-CRISPR/Cas12a reactions with 48 clinical samples and compare the results with those of PCR-Fluorescent Probe Method in clinical applications and the qPCR method for detecting the UL123 gene.

Results

The optimized RPA-CRISPR system exhibited high sensitivity and specificity for HCMV detection. The positive rate of clinical sample detection was approximately 20.5 % (6/48) higher than that of the clinical detection method.

Conclusion

Currently, the sensitivity and early detection of HCMV in clinical settings are still limited. The UL123 gene of HCMV is characterized by high transcription in the early stage and high conservation. The RPA-CRISPR/Cas12a technology exhibits high sensitivity in detecting the HCMV UL123 gene, and it is expected to provide a more effective method for the early specific detection of HCMV infection.

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基于HCMV-UL123基因的RPA-CRISPR/Cas12a检测：一种比临床检测方法检出率更高的方法

目的人巨细胞病毒（HCMV）是一种流行的双链DNA包膜病毒，对免疫功能低下的个体构成威胁。目前的临床检测方法灵敏度不足，需要更有效的方法。方法基于HCMV UL123基因设计筛选RPA引物和crRNA。利用克隆质粒和hcmv感染细胞的全基因组样本评价HCMV-RPA-CRISPR检测方法。对48份临床样本进行RPA-CRISPR/Cas12a反应，并与临床应用的pcr -荧光探针法和qPCR检测UL123基因的结果进行比较。结果优化后的RPA-CRISPR系统检测HCMV具有较高的灵敏度和特异性。临床样品检测阳性率比临床检测方法高20.5 %（6/48）。结论目前临床对HCMV的敏感性和早期检测仍然有限。HCMV UL123基因具有早期高转录和高保守性的特点。RPA-CRISPR/Cas12a技术对HCMV UL123基因的检测具有较高的敏感性，有望为HCMV感染的早期特异性检测提供更有效的方法。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of Infection and Public Health PUBLIC, ENVIRONMENTAL & OCCUPATIONAL HEALTH -INFECTIOUS DISEASES

CiteScore

13.10

自引率

1.50%

发文量

203

审稿时长

96 days

期刊介绍： The Journal of Infection and Public Health, first official journal of the Saudi Arabian Ministry of National Guard Health Affairs, King Saud Bin Abdulaziz University for Health Sciences and the Saudi Association for Public Health, aims to be the foremost scientific, peer-reviewed journal encompassing infection prevention and control, microbiology, infectious diseases, public health and the application of healthcare epidemiology to the evaluation of health outcomes. The point of view of the journal is that infection and public health are closely intertwined and that advances in one area will have positive consequences on the other. The journal will be useful to all health professionals who are partners in the management of patients with communicable diseases, keeping them up to date. The journal is proud to have an international and diverse editorial board that will assist and facilitate the publication of articles that reflect a global view on infection control and public health, as well as emphasizing our focus on supporting the needs of public health practitioners. It is our aim to improve healthcare by reducing risk of infection and related adverse outcomes by critical review, selection, and dissemination of new and relevant information in the field of infection control, public health and infectious diseases in all healthcare settings and the community.