A Simplified Amplification‐Free Strategy with Lyophilized CRISPR‐CcrRNA System for Drug‐Resistant Salmonella Detection

Abstract

With the widespread abuse of antibiotics, the problem of bacterial resistance is becoming increasingly serious, resulting in more than 700 000 deaths annually, worldwide. [1] Salmonella, one of the most dangerous and infectious foodborne pathogens, ranks second among pathogens that endanger public food safety. [2] Salmonella accounts for 41% of global diarrhea-related deaths. [1,3] The emergence of drug-resistant Salmonella has increased the danger to public health. Early diagnosis and prompt detection of drug resistance require a rapid, sensitive, and cost-effective detection program from the perspective of prevention, screening for drug-resistant bacteria, and management of food-related accidents. [4,5] Traditional culture-based antibiotic sensitivity testing (AST) methods are complicated and often require 2 to 7 days. [6] In contrast, nucleic acid detection of drug-resistance genes has significant advantages in terms of sensitivity and specificity. [7] The gold standard for nucleic acid detection, quantitative polymerase chain reaction (qPCR), can detect 10 CFU mL −1 of resistant bacteria within a few hours. [8,9] However, it still relies heavily on specialized equipment and expertise and is difficult to use outside a laboratory setting. [4] Lateral flow assay (LFA) is the most promising point-of-care testing (POCT) platform owing to its minimal device dependence, speed, sensitivity, and user-friendliness. [10,11] A visual readout LFA based on antigen capture with a faster and more accurate reading within 15 min was developed. [12] Unfortunately, this immunology-based pathogenic bacterial detection via LFA generally has low sensitivity and cannot be used for AST. [10,13] Combining the CRISPR/Cas system with LFA allows for sensitive and specific detection of targets while maintaining its fast and user-friendly features. For instance. Gootenberg et al [14] proposed a method called SHERLOCK for SARS-CoV-2 detection, [15] Joung et al. [13] demonstrated the direct detection of SARS-CoV-2 samples and further developed the SHINE. [16] All of the LFA-CRISPR methods mentioned above enable Drug resistance in pathogenic bacteria has become a major threat to global health. The misuse of antibiotics has increased the number of resistant bacteria in the absence of rapid, accurate, and cost-effective diagnostic tools. Here, an amplification-free CRISPR-Cas12a time-resolved fluorescence immunochromatographic assay (AFC-TRFIA) is used to detect drugresistant Salmonella. Multi-locus targeting in combination crRNA (CcrRNA) is 27-fold more sensitive than a standalone crRNA system. The lyophilized CRISPR system further simplifies the operation and enables one-pot detection. Induction of nucleic acid fixation via differentially charged interactions reduced the time and cost required for flowmetric chromatography with enhanced stability. The induction of nucleic acid fixation via differentially charged interactions reduces the time and cost required for flowmetric chromatography with enhanced stability. The platform developed for th...

Palm‐Sized Lab‐In‐A‐Magnetofluidic Tube Platform for Rapid and Sensitive Virus Detection

“…In the future, we will integrate an automatic magnetofluidic operation into the processor to enhance the automation of the LIAMT assay [ 43 ] . In addition, we will further lyophilize the reagents in the LIAMT device to simplify the device operation [ 44 , 45 ] . Overall, our simple, portable, sensitive, and self‐contained LIAMT platform is a promising tool for rapid detection of infectious disease at the point‐of‐care, especially in resource‐limited settings.…”

High-Performance Fluorescent Microspheres Based on Fluorescence Resonance Energy Transfer Mode for Lateral Flow Immunoassays

“…8 However, the traditional LFIA with a colorimetric readout (colloidal gold) confronted some shortcomings in terms of qualitative results, insufficient sensitivity, and poor accuracy. 9 Therefore, fluorescent materials with excellent optical properties, such as carbon dots, 10 quantum dot beads (QBs), 11 and time-resolved fluorescent microspheres, 12 had been introduced into LFIA for improving the sensitivity in a quantitative way. 13 However, these fluorescent materials suffered from the aggregationinduced quenching (ACQ) effect, 14 resulting in low fluorescence signals in the aggregated state owing to intermolecular π−π interactions or other nonradiative channels to form radical conjugates.…”