A novel CRISPR-Cas12a assay for African swine fever virus achieved 98.3% sensitivity and 100% specificity in whole blood samples, with visual results available in as few as 7 minutes.
Does an RPA-dependent CRISPR-Cas12a assay accurately detect ASFV in diagnostic samples compared to rtPCR?
A novel CRISPR-Cas12a assay provides rapid, highly sensitive, and specific point-of-care detection of African swine fever virus, comparable to rtPCR.
Absolute Event Rate: 0% vs 0%
African swine fever virus (ASFV) is a highly transmissible pathogen affecting swine, causing a devastating disease with high mortality rates in naive populations. Given the likelihood of significant economic impacts associated with an ASF outbreak, considerable resources have been allocated in the United States to safeguard the swine industry against this threat. Ongoing outbreaks of ASF in the Dominican Republic and Haiti further threaten the U.S. swine industry, given their proximity and involvement in movement to and from North America. Although surveillance programs are ongoing, limited point-of-care (POC) tests are available during outbreaks with the sensitivity and specificity standards of laboratory testing (e.g., real-time PCR rtPCR). However, the recently developed CRISPR-Cas-based testing systems may offer comparable high-quality results. We sought to develop a low-cost visual detection method for ASFV by employing a recombinase polymerase amplification (RPA)-dependent CRISPR-Cas12a technique that can be utilized in the field as a POC assay. Our CRISPR-Cas12a assay had comparable sensitivity and specificity to rtPCR, both visually and when quantified using a fluorescence reader. In whole blood samples from ASFV-suspect or ASFV-negative cases, our CRISPR assay achieved a sensitivity of 98.3% (102 DNA copies) and a specificity of 100%. Test results of our RPA-CRISPR assay can be visualized in as few as 7 min, with peak fluorescence at 40 min (RPA and CRISPR steps). Our results lay the groundwork for a large-scale POC assay assessment for ASFV detection and offer a robust workflow that works with commonly submitted diagnostic samples.
Kambakam et al. (Wed,) reported a other. A novel CRISPR-Cas12a assay for African swine fever virus achieved 98.3% sensitivity and 100% specificity in whole blood samples, with visual results available in as few as 7 minutes.