Abstract
Bacteria have been extensively utilized as living therapeutics for disease treatment due to their unique characteristics, such as genetic manipulability, rapid proliferation and specificity to target disease sites. Various in vivo insults can, however, decrease the vitality of dosed bacteria, leading to low overall bioavailability. Additionally, the innate antigens on the bacterial surface and the released toxins and metabolites may cause undesired safety issues. These limitations inevitably result in inadequate treatment outcomes, thereby hindering the clinical transformation of living bacterial therapeutics. Recently, we have developed a versatile platform to prepare advanced living bacterial therapeutics by nanocoating bacteria individually via either chemical decoration or physical encapsulation, which can improve bioavailability and reduce side effects for enhanced microbial therapy. Here we use interfacial self-assembly to prepare lipid membrane-coated bacteria (LCB), exhibiting increased resistance against a variety of harsh environmental conditions owing to the nanocoating’s protective capability. Meanwhile, we apply mechanical extrusion to generate cell membrane-coated bacteria (CMCB), displaying improved biocompatibility owing to the nanocoating’s shielding effect. We describe their detailed preparation procedures and demonstrate the expected functions of the coated bacteria. We also show that following oral delivery and intravenous injection in mouse models, LCB and CMCB present appealing potential for treating colitis and tumors, respectively. Compared with bioengineering that lacks versatile molecular tools for heterogeneous expression, the surface nanocoating technique is convenient to introduce functional components without restriction on bacterial strain types. Excluding bacterial culture, the fabrication of LCB takes ~2 h, while the preparation of CMCB takes ~5 h.
Key points
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This protocol adds a surface nanocoating to bacteria via either chemical decoration or physical encapsulation to improve bioavailability and reduce side effects for enhanced microbial therapy.
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Compared with biological engineering or genetic modification, surface nanocoating can easily introduce various functional components and can be applied to diverse bacterial strains. It can be achieved using either native or Food and Drug Administration-approved synthetic materials, ensuring satisfactory biocompatibility and safety.
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Data availability
The main data discussed in this protocol are available in the supporting primary research papers (refs. 21,39). Source data are provided with this paper.
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Acknowledgements
This work was financially supported by the National Key Research and Development Program of China (2021YFA0909400 to J.L.), the National Natural Science Foundation of China (32101218 to Z.C.), the Explorer Program of the Science and Technology Commission of Shanghai Municipality (21TS1400400 to J.L.), the Innovative Research Team of High-Level Local Universities in Shanghai (SHSMU-ZDCX20210900 to J.L.), the Foundation of National Infrastructures for Translational Medicine (Shanghai) (TMSK-2021-123 to J.L.), and the Two-hundred Talent (20181704 to J.L.).
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J.L. and Z.C. originated the methods of preparing LCB and CMCB. All authors wrote the manuscript and approved the contents of the protocol.
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J.L. and Z.C. are inventors on a filed provisional application China patent no. CN201911034435.7 (Surface modified bacteria, preparation method and application thereof), submitted by Shanghai Jiao Tong University School of Medicine Affiliated Renji Hospital that covers the potential therapeutic uses of coated bacteria for treating colitis and cancer.
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The authors declare no competing interests.
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Nature Protocols thanks Fu-Gen Wu and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
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Key references using this protocol
Cao, Z. et al. Nat. Commun. 10, 5783 (2019): https://doi.org/10.1038/s41467-019-13727-9
Geng, Z. et al. Sci. Adv. 9, eade0997 (2023): https://doi.org/10.1126/sciadv.ade0997
Cao, Z. et al. Nat. Commun. 10, 3452 (2019): https://doi.org/10.1038/s41467-019-11390-8
Lin, S. et al. Sci. Adv. 7, eabf0677 (2021): https://doi.org/10.1126/sciadv.abf0677
Wang, X. et al. Sci. Adv. 6, eabb1952 (2020): https://doi.org/10.1126/sciadv.abb1952
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Cao, Z., Liu, J. Surface nanocoating of bacteria as a versatile platform to develop living therapeutics. Nat Protoc (2024). https://doi.org/10.1038/s41596-024-01019-6
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DOI: https://doi.org/10.1038/s41596-024-01019-6
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