Infection and Drug Resistance (Feb 2022)

A New Method Based on LAMP-CRISPR–Cas12a-Lateral Flow Immunochromatographic Strip for Detection

  • Xu H,
  • Tang H,
  • Li R,
  • Xia Z,
  • Yang W,
  • Zhu Y,
  • Liu Z,
  • Lu G,
  • Ni S,
  • Shen J

Journal volume & issue
Vol. Volume 15
pp. 685 – 696

Abstract

Read online

Huaming Xu,1 Hao Tang,2 Rongrong Li,3 Zhaoxin Xia,2 Wensu Yang,2 Yi Zhu,2 Zhen Liu,2 Guoping Lu,4 Shenwang Ni,2 Jilu Shen1 1Department of Laboratory Medicine, The Fourth Affiliated Hospital of Anhui Medical University, Hefei, 230012, People’s Republic of China; 2University Laboratory, The Fourth Affiliated Hospital of Anhui Medical University, Hefei, 230012, People’s Republic of China; 3The First Affiliated Hospital of Anhui Medical University Laboratory Department, Hefei, Anhui, Peoples’ Republic of China; 4Laboratory Department of Fuyang Hospital Affiliated to Anhui Medical University, Fuyang, Anhui, People’s Republic of ChinaCorrespondence: Jilu Shen, Tel +86 151 5515 2963, Email [email protected]; [email protected]: Carbapenemase-mediated antimicrobial resistance is currently a hot spot of global concern. Carbapenem-resistant organisms are highly prevalent in hospitals associated with difficult-to-treat infections, resulting in poor clinical outcome due to limited treatment options. It is urgently needed to have a rapid, efficient, and convenient molecular assay for identifying such resistant strains.Methods: For this end, we developed a new laboratory assay targeting Klebsiella pneumoniae carbapenemase (KPC) and New Delhi metallo-β-lactamase (NDM) based on loop-mediated isothermal amplification, CRISPR–Cas12a, and lateral flow immunochromatographic strip (CRISPR–Cas-LAMP-lateral flow strip). The method was designed to use a guide RNA (gRNA) to recognize the target DNA and guide Cas12a to cleave the target DNA, and simultaneously cleave any single-stranded DNA within the cleavage reaction system.Results: The cleavage products are visible to the naked eye on the lateral flow strip. This method is highly sensitive in direct detection of bacteria in samples containing at least 3× 105 CFU/mL without the need for bacterial culture.Discussion: It provides shorter turnaround time and higher specificity than the conventional bacterial culture and susceptibility testing method. This new assay is applicable for extensive use in hospital infection control, as well as identification and treatment of resistant strains due to simple operation and inexpensive apparatuses.Keywords: carbapenemase, Klebsiella pneumoniae carbapenemase, New Delhi metallo-β-lactamase, loop-mediated isothermal amplification, CRISPR–Cas12a, lateral flow immunochromatographic strip, detection

Keywords