AUTHOR=Zhou Zihan , Liang Lina , Liao Chuan , Pan Lele , Wang Chunfang , Ma Jiangmei , Yi Xueli , Tan Meiying , Li Xuebin , Wei Guijiang 

TITLE=A multiplex RPA coupled with CRISPR-Cas12a system for rapid and cost-effective identification of carbapenem-resistant Acinetobacter baumannii

JOURNAL=Frontiers in Microbiology

VOLUME=15

YEAR=2024

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2024.1359976

DOI=10.3389/fmicb.2024.1359976

ISSN=1664-302X

ABSTRACT=<sec><title>Background</title><p>Carbapenem-resistant <italic>Acinetobacter baumannii</italic> (CRAB) poses a severe nosocomial threat, prompting a need for efficient detection methods. Traditional approaches, such as bacterial culture and PCR, are time-consuming and cumbersome. The CRISPR-based gene editing system offered a potential approach for point-of-care testing of CRAB.</p></sec><sec><title>Methods</title><p>We integrated recombinase polymerase amplification (RPA) and CRISPR-Cas12a system to swiftly diagnose CRAB-associated genes, <italic>OXA-51</italic> and <italic>OXA-23</italic>. This multiplex RPA-CRISPR-Cas12a system eliminates bulky instruments, ensuring a simplified UV lamp-based outcome interpretation.</p></sec><sec><title>Results</title><p>Operating at 37°C to 40°C, the entire process achieves CRAB diagnosis within 90 minutes. Detection limits for <italic>OXA-51</italic> and <italic>OXA-23</italic> genes are 1.3 × 10<sup>−6</sup> ng/μL, exhibiting exclusive CRAB detection without cross-reactivity to common pathogens. Notably, the platform shows 100% concordance with PCR when testing 30 clinical <italic>Acinetobacter baumannii</italic> strains.</p></sec><sec><title>Conclusion</title><p>In conclusion, our multiplex RPA coupled with the CRISPR-Cas12a system provides a fast and sensitive CRAB detection method, overcoming limitations of traditional approaches and holding promise for efficient point-of-care testing.</p></sec>