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Automatic Microfluidic Harmonized RAA-CRISPR Diagnostic System for Rapid and Accurate Identification of Bacterial
Xinran Xiang1,2, Xiaoqing Ren3, Qianyu Wen4
1Fujian Key Laboratory of Aptamers Technology, Fuzhou General Clinical Medical School (the 900th Hospital), Fujian Medical University, Fuzhou 350001, China.
Abstract:
Respiratory tract infections (RTIs) pose a grave threat to human health, with bacterial pathogens being the primary culprits behind severe illness and mortality. In response to the pressing issue, we developed a centrifugal microfluidic chip integrated with a recombinase-aided amplification (RAA)-clustered regularly interspaced short palindromic repeats (CRISPR) system to achieve rapid detection of respiratory pathogens. The limitations of conventional two-step CRISPR-mediated systems were effectively addressed by employing the all-in-one RAA-CRISPR detection method, thereby enhancing the accuracy and sensitivity of bacterial detection. Moreover, the integration of a centrifugal microfluidic chip led to reduced sample consumption and significantly improved the detection throughput, enabling the simultaneous detection of multiple respiratory pathogens. Furthermore, the incorporation of Chelex-100 in the sample pretreatment enabled a sample-to-answer capability. This pivotal addition facilitated the deployment of the system in real clinical sample testing, enabling the accurate detection of 12 common respiratory bacteria within a set of 60 clinical samples. The system offers rapid and reliable results that are crucial for clinical diagnosis, enabling healthcare professionals to administer timely and accurate treatment interventions to patients.
Insights
This study presents a rapid, all-in-one RAA-CRISPR diagnostic system on a microfluidic chip for detecting respiratory pathogens. It enables quick and accurate identification of 12 common bacteria from clinical samples.
Area of Science:
- Biotechnology
- Microbiology
- Medical Diagnostics
Background:
- Bacterial respiratory tract infections (RTIs) are a major global health concern, causing significant morbidity and mortality.
- Current diagnostic methods for respiratory pathogens often lack the speed and sensitivity required for timely clinical intervention.
Purpose of the Study:
- To develop a rapid, sensitive, and high-throughput diagnostic system for the simultaneous detection of multiple respiratory bacterial pathogens.
- To overcome the limitations of conventional two-step CRISPR-mediated detection systems.
Main Methods:
- Integration of a recombinase-aided amplification (RAA) with a clustered regularly interspaced short palindromic repeats (CRISPR) system on a centrifugal microfluidic chip.
- Development of an all-in-one RAA-CRISPR assay for enhanced bacterial detection.
- Inclusion of Chelex-100 for simplified sample pretreatment, enabling a sample-to-answer workflow.
Main Results:
- The RAA-CRISPR system demonstrated enhanced accuracy and sensitivity compared to conventional methods.
- The microfluidic chip facilitated reduced sample consumption and increased detection throughput for simultaneous pathogen analysis.
- The system successfully detected 12 common respiratory bacteria in 60 clinical samples with high accuracy.
Conclusions:
- The developed centrifugal microfluidic RAA-CRISPR system provides a rapid, reliable, and efficient platform for diagnosing bacterial respiratory infections.
- This technology has the potential to significantly improve clinical decision-making and patient treatment outcomes by enabling timely diagnosis.
