D-CAPS: an efficient CRISPR-Cas9-based phage defense system for E. coli

Mingjun Sun, Jie Gao, Hongjie Tang, Hengyi Wang, Liyan Zhou, Chuan Song, Yongqiang Tian, and Qi Li

During industrial microbial fermentation, bacteriophage contamination is a significant risk factor leading to production failure and reduced yields. T7 phage, a representative phage with a short lytic cycle and strong virulence, often causes acute lysis of Escherichia coli, severely threatening the stability of fermentation systems. This study constructed an inducible-free anti-phage system based on the CRISPR/Cas9 system and demonstrated that this system achieved complete defense against T7 phage in E. coli. Even at an infection multiplicity of 2, it maintained normal growth of the host cells and preserved their ability to express the target protein normally. As depicted in the cover illustration: the scorpion represents the Cas9 protein protecting the bacterium against phage invasion; the two intertwined snakes represent the invading phage DNA; the scorpion’s tail represents the gRNA——its entanglement with the snakes symbolizes the recognition of the target DNA by the gRNA; the scorpion’s claws shearing the snake’s body represent the Cas9 enzyme’s targeted recognition and cleavage of the DNA. By simultaneously targeting two essential genes of T7, this system effectively overcomes the escape problem inherent in single-target systems. This research provides an efficient and eco-friendly new strategy for phage defense in industrial fermentation strains.

This cover is designed by Yinxi Zhou and Jianfeng Jin from Hainan Medical University.