Application of CRISPR-Cas system in the diagnosis and therapy of ESKAPE infections

Abstract

Antimicrobial-resistant ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens represent a global threat to human health. ESKAPE pathogens are the most common opportunistic pathogens in nosocomial infections, and a considerable number of their clinical isolates are not susceptible to conventional antimicrobial therapy. Therefore, innovative therapeutic strategies that can effectively deal with ESKAPE pathogens will bring huge social and economic benefits and ease the suffering of tens of thousands of patients. Among these strategies, CRISPR (clustered regularly interspaced short palindromic repeats) system has received extra attention due to its high specificity. Regrettably, there is currently no direct CRISPR-system-based anti-infective treatment. This paper reviews the applications of CRISPR-Cas system in the study of ESKAPE pathogens, aiming to provide directions for the research of ideal new drugs and provide a reference for solving a series of problems caused by multidrug-resistant bacteria (MDR) in the post-antibiotic era. However, most research is still far from clinical application.

Keywords: CRISPR-Cas; ESKAPE; diagnosis; infection; pathogen; therapy.

Figure 1

Composition and function of CRISPR-Cas system. CRISPR/Cas system is an adaptive immune defense formed by bacteria and archaea in the long-term evolution process, which fights against invading viruses and foreign DNA through three different but continuous stages: adaptation, crRNA maturation, and interference. CRISPR-Cas system mainly includes two categories: category 1 and category 2. Type 1 CRISPR-Cas systems include types I, III, and IV, and type 2 systems include types II, V, and VI. The first type of CRISPR-Cas system uses the cooperation of multiple Cas proteins to interfere, while the second type of system is interfered by a single protein.

Figure 2

The second type of system is interfered by a single protein. Basic strategy of using exogenous CRISPR-Cas as antibacterial agent. CRISPR-Cas phage vector is a plasmid construct cloned with CRISPR-Cas element and phage packaging sequence. Phage carrier particles are adsorbed on bacteria, and plasmids carrying genes related to CRISPR-Cas system are transferred to drug-resistant bacteria. Exogenous Cas gene and sgRNA expression sequence were expressed in host cells. The cleavage of targeted drug-resistant genes by CRISPR-Cas system will lead to the loss of drug-resistant plasmids or DNA double-strand breaks. When bacteria cannot re-cyclize plasmids or repair their genomes, drug-resistant bacteria will die. Phage continues to enter the replication cycle and synthesize offspring and lyse cells, further optimizing the therapeutic effect.