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Updated: Jul 10, 2025
In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
Published on: March 25, 2020
Appelmans protocol - A directed in vitro evolution enables induction and recombination of prophages with expanded
Thao Nguyen Vu1, Justin Ryan Clark2, Eris Jang3
1Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Graduate School of Medical Science, Brain Korea 21 Project, Yonsei University College of Medicine, Seoul, South Korea.
Abstract:
Infections caused by carbapenem-resistant Acinetobacter baumannii (CRAB) present significant healthcare challenges due to limited treatment options. Bacteriophage (phage) therapy offers potential as an alternative treatment. However, the high host specificity of phages poses challenges for their therapeutic application. To broaden the phage spectrum, laboratory-based phage training using the Appelmans protocol was employed in this study. As a result, the protocol successfully expanded the host range of a phage cocktail targeting CRAB. Further analysis revealed that the expanded host range phages isolated from the output cocktail were identified as recombinant derivatives originating from prophages induced from encountered bacterial strains. These findings provide valuable genetic insights into the protocol's mechanism when applied to phages infecting A. baumannii strains that have never been investigated before. However, it is noteworthy that the expanded host range phages obtained from this protocol exhibited limited stability, raising concerns about their suitability for therapeutic purposes.
