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Table 1 Successful examples of designed CRISPR-Cas antimicrobials and delivery strategies

From: Engineered CRISPR-Cas systems for the detection and control of antibiotic-resistant infections

Delivery systems

Bacteria

CRISPR-Cas locus

Brief Result

Refs.

Phage

E. coli

λCas-CRISPR

Bacteria containing CRISPR-Cas targeting blaCTX-M-15 and blaNDM-1 genes had low transformation efficiency of AMR plasmids carrying these genes

[68]

S. aureus

CRISPR-Cas9

A phagemid pDB91 targeting mecA was designed and encapsulated in phage ΦNM1, and MRSA was resensitized to methicillin

[60]

E. coli

CRISPR-Cas9

RNA-guided nucleases induced double-strand breaks in blaSHV-18 or blaNDM-1, which reduced the transduction efficiency of plasmids containing these genes by approximately 1000-fold

[59]

E.coli

CRISPR-Cas13a

CRISPR-Cas13a-induced bacteria decreased by approximately three orders and demonstrated sequence-specific killing activity against bacteria carrying the blaIMP-1 gene in an EC-CapsidCas13a_blaIMP-1 concentration-dependent manner

[85]

S. aureus

CRISPR-Cas9

Integration of CRISPR-Cas9 system into Ï•SaBov temperate phage genome, removal of nuc gene from the host chromosome, and expansion of host specificity of the phage was achieved by complementing the tail fibre protein

[64]

C. difficile

CRISPR-Cas3, Type I-B CRISPR-Cas system

The endogenous type I-B CRISPR-Cas system in C. difficile redirects endogenous CRISPR-Cas3 activity against the bacterial chromosome. A recombinant bacteriophage expressing bacterial genome-targeting CRISPR RNAs was significantly more effective than its wild-type parent bacteriophage at killing C. difficile

[66]

Mobile genomic island

S. aureus

CRISPR-dCas9, CRISPR-Cas9

Highly mobile SAPIs were used to treat S. aureus infections, and ABD2003 killed S. aureus by introducing double-strand breaks in the agrA loci of the chromosome

[109]

Conjugative plasmid

E. coli

Types I-E CRISPR-Cas system

E. coli K-12 and B strains were removed by targeting fucP gene and ogr gene, respectively, and the two strains were removed by targeting the groL gene, demonstrating the sequence-specific removal function of CRISPR-Cas

[63]

E. coli, Salmonella

CRISPR-Cas9

Plasmids based on the IncP RK2 conjugative system can be used as delivery vectors for a TevSpCas9 dual nuclease. Targeting of single or multiplexed sgRNAs to non-essential genes resulted in high S. enterica killing efficiencies

[110]

E. coli

CRISPR-Cas9

An innovative strategy based on targeted-antibacterial-plasmids (TAPs) that uses bacterial conjugation to deliver CRISPR-Cas systems exerting a strain-specific antibacterial activity. TAPs directed against a plasmid-borne carbapenem resistance gene efficiently resensitized the strain to the drug

[114]

E. coli

CRISPR-Cas9

The conjugative plasmid was used to deliver the CRISPR-Cas9 system targeting the mcr-1 gene, restoring sensitivity to polymyxin in E. coli

[82]

Conjugative plasmid

E. coli

CRISPR-Cas9

The pMob-Cas9 plasmid carrying the CRISPR-Cas9 system was conjugated to E. coli for targeted clearance of the mcr-1 gene

[81]

S. algae

CRISPR-Cas9

CRISPR-Cas9 is used to target the sul2, blaOXA-55-like, and NmcR-like genes, making S. algae less resistant to carbapenem antibiotics

[78]

E. coli

CRISPR-Cas9

sopA, nikA, mcr-1, vagC, and hicB antitoxin genes were used as target genes for the clearance of drug-resistant plasmids

[73]

E. faecalis

CRISPR-Cas9

Description of the adaption of type II CRISPR-Cas system encoded on a pheromone-responsive conjugative plasmid that was efficiently transferred to E. faecalis for the selective removal of ermB and tetM

[75]

Hydrogel

S. aureus

CRISPR-Cas9

Quantitative antibiofilm effects increased over time for Fosfomycin-phage (dual) therapeutics delivered via alginate hydrogel. This module was successfully used to reduce soft tissue infection but not bone infection

[196]

Electroporation

S. aureus

CRISPR-dCas9

Electroporation technology was used to deliver CRISPR-dCas9 into S. aureus, inducing downregulation of tarH, tarO, and tarG genes and making the bacteria sensitive to lysostaphin

[77]

Nanoparticle

S. aureus

CRISPR-Cas9

The transfection efficiency of MRSA was significantly improved by mixing SpCas9-bPEI with sgRNA to form a nanosized CRISPR complexes (= Cr-Nanocomplex) designed to target mecA, which is a major gene associated with methicillin resistance

[67]