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Hydrogel-based biocontainment of bacteria for continuous sensing and computation

Nature Chemical Biology volume 17, pages 724–731 (2021)Cite this article

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Abstract

Genetically modified microorganisms (GMMs) can enable a wide range of important applications including environmental sensing and responsive engineered living materials. However, containment of GMMs to prevent environmental escape and satisfy regulatory requirements is a bottleneck for real-world use. While current biochemical strategies restrict unwanted growth of GMMs in the environment, there is a need for deployable physical containment technologies to achieve redundant, multi-layered and robust containment. We developed a hydrogel-based encapsulation system that incorporates a biocompatible multilayer tough shell and an alginate-based core. This deployable physical containment strategy (DEPCOS) allows no detectable GMM escape, bacteria to be protected against environmental insults including antibiotics and low pH, controllable lifespan and easy retrieval of genomically recoded bacteria. To highlight the versatility of DEPCOS, we demonstrated that robustly encapsulated cells can execute useful functions, including performing cell–cell communication with other encapsulated bacteria and sensing heavy metals in water samples from the Charles River.

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Fig. 1: Schematic of the DEPCOS platform.
Fig. 2: Cell encapsulation in tough hydrogel capsules.
Fig. 3: Tough hydrogel shell provides robust biocontainment.
Fig. 4: Combining chemical and physical containment strategies for optimal biocontainment and protection.
Fig. 5: Sensing, recording and communication capabilities of encapsulated bacterial cells.

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Data availability

Data supporting this study are presented in the main text and Supplementary Information, and are available from the corresponding authors upon request. Source data are provided with this paper.

References

  1. Singh, J. S., Abhilash, P. C., Singh, H. B., Singh, R. P. & Singh, D. P. Genetically engineered bacteria: an emerging tool for environmental remediation and future research perspectives. Gene 480, 1–9 (2011).

    Article  CAS  PubMed  Google Scholar 

  2. Farrar, K., Bryant, D. & Cope-Selby, N. Understanding and engineering beneficial plant-microbe interactions: plant growth promotion in energy crops. Plant Biotechnol. J. 12, 1193–1206 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  3. Tang, T.-C. et al. Materials design by synthetic biology. Nat. Rev. Mater. https://doi.org/10.1038/s41578-020-00265-w (2020).

  4. Dana, G. V., Kuiken, T., Rejeski, D. & Snow, A. A. Synthetic biology: four steps to avoid a synthetic-biology disaster. Nature 483, 29 (2012).

    Article  CAS  PubMed  Google Scholar 

  5. Torres, L., Krüger, A., Csibra, E., Gianni, E. & Pinheiro, V. B. Synthetic biology approaches to biological containment: pre-emptively tackling potential risks. Essays Biochem. 60, 393–410 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  6. Epstein, M. M. & Vermeire, T. Scientific opinion on risk assessment of synthetic biology. Trends Biotechnol. 34, 601–603 (2016).

    Article  CAS  PubMed  Google Scholar 

  7. Wright, O., Stan, G. B. & Ellis, T. Building-in biosafety for synthetic biology. Microbiol. 159, 1221–1235 (2013).

    Article  CAS  Google Scholar 

  8. Lee, J. W., Chan, C. T. Y., Slomovic, S. & Collins, J. J. Next-generation biocontainment systems for engineered organisms. Nat. Chem. Biol. 14, 530–537 (2018).

    Article  CAS  PubMed  Google Scholar 

  9. Chan, C. T. Y., Lee, J. W., Cameron, D. E., Bashor, C. J. & Collins, J. J. ‘Deadman’ and ‘Passcode’ microbial kill switches for bacterial containment. Nat. Chem. Biol. 12, 82–86 (2016).

    Article  CAS  PubMed  Google Scholar 

  10. Wright, O., Delmans, M., Stan, G. B. & Ellis, T. GeneGuard: a modular plasmid system designed for biosafety. ACS Synth. Biol. 4, 307–316 (2015).

    Article  CAS  PubMed  Google Scholar 

  11. Gallagher, R. R., Patel, J. R., Interiano, A. L., Rovner, A. J. & Isaacs, F. J. Multilayered genetic safeguards limit growth of microorganisms to defined environments. Nucleic Acids Res. 43, 1945–1954 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Rovner, A. J. et al. Recoded organisms engineered to depend on synthetic amino acids. Nature 518, 89–93 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Hoffman, A. S. Hydrogels for biomedical applications. Adv. Drug Deliv. Rev. 64, 18–23 (2012).

    Article  Google Scholar 

  14. Choi, M. et al. Light-guiding hydrogels for cell-based sensing and optogenetic synthesis in vivo. Nat. Photonics 7, 987–994 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Anselmo, A. C., McHugh, K. J., Webster, J., Langer, R. & Jaklenec, A. Layer-by-layer encapsulation of probiotics for delivery to the microbiome. Adv. Mater. 28, 9486–9490 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Lee, K. Y. & Mooney, D. J. Alginate: properties and biomedical applications. Prog. Polym. Sci. 37, 106–126 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Kearney, C. J. & Mooney, D. J. Macroscale delivery systems for molecular and cellular payloads. Nat. Mater. 12, 1004–1017 (2013).

    Article  CAS  PubMed  Google Scholar 

  18. Billiet, T., Vandenhaute, M., Schelfhout, J., Van Vlierberghe, S. & Dubruel, P. A review of trends and limitations in hydrogel-rapid prototyping for tissue engineering. Biomaterials 33, 6020–6041 (2012).

    Article  CAS  PubMed  Google Scholar 

  19. Kim, B. J. et al. Cytoprotective alginate/polydopamine core/shell microcapsules in microbial encapsulation. Angew. Chem. Int. Ed. 53, 14443–14446 (2014).

    Article  CAS  Google Scholar 

  20. Li, P., Müller, M., Chang, M. W., Frettlöh, M. & Schönherr, H. Encapsulation of autoinducer sensing reporter bacteria in reinforced alginate-based microbeads. ACS Appl. Mater. Interfaces 9, 22321–22331 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Zhang, B. B., Wang, L., Charles, V., Rooke, J. C. & Su, B. L. Robust and biocompatible hybrid matrix with controllable permeability for microalgae encapsulation. ACS Appl. Mater. Interfaces 8, 8939–8946 (2016).

    Article  CAS  PubMed  Google Scholar 

  22. Sun, J. Y. et al. Highly stretchable and tough hydrogels. Nature 489, 133–136 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Liu, X. et al. Stretchable living materials and devices with hydrogel-elastomer hybrids hosting programmed cells. Proc. Natl Acad. Sci. USA 114, 2200–2205 (2017).

    Article  CAS  PubMed  Google Scholar 

  24. Valade, D., Wong, L. K., Jeon, Y., Jia, Z. & Monteiro, M. J. Polyacrylamide hydrogel membranes with controlled pore sizes. J. Polym. Sci., Part A: Polym. Chem. 51, 129–138 (2013).

    Article  CAS  Google Scholar 

  25. Atkinson, J. The Mechanics of Soils and Foundations 2nd edn (CRC Press, 2007); https://doi.org/10.1201/9781315273549

  26. Liu, X. et al. Ingestible hydrogel device. Nat. Commun. 10, 493 (2019).

  27. Houghton, L. A. et al. Motor activity of the gastric antrum, pylorus, and duodenum under fasted conditions and after a liquid meal. Gastroenterology 94, 1276–1284 (1988).

    Article  CAS  PubMed  Google Scholar 

  28. Zarket, B. C. & Raghavan, S. R. Onion-like multilayered polymer capsules synthesized by a bioinspired inside-out technique. Nat. Commun. 8, 193 (2017).

  29. Eun, Y. J., Utada, A. S., Copeland, M. F., Takeuchi, S. & Weibel, D. B. Encapsulating bacteria in agarose microparticles using microfluidics for high-throughput cell analysis and isolation. ACS Chem. Biol. 6, 260–266 (2011).

    Article  CAS  PubMed  Google Scholar 

  30. Kong, H. J., Kim, E. S., Huang, Y. C. & Mooney, D. J. Design of biodegradable hydrogel for the local and sustained delivery of angiogenic plasmid DNA. Pharm. Res. 25, 1230–1238 (2008).

    Article  CAS  PubMed  Google Scholar 

  31. Farzadfard, F. & Lu, T. K. Genomically encoded analog memory with precise in vivo DNA writing in living cell populations. Science 346, 1256272–1256272 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  32. Golmohamadi, M. & Wilkinson, K. J. Diffusion of ions in a calcium alginate hydrogel-structure is the primary factor controlling diffusion. Carbohydr. Polym. 94, 82–87 (2013).

    Article  CAS  PubMed  Google Scholar 

  33. Li, Z. et al. Biofilm-inspired encapsulation of probiotics for the treatment of complex infections. Adv. Mater. 30, e1803925 (2018).

    Article  PubMed  Google Scholar 

  34. Bjerketorp, J., Håkansson, S., Belkin, S. & Jansson, J. K. Advances in preservation methods: keeping biosensor microorganisms alive and active. Curr. Opin. Biotechnol. 17, 43–49 (2006).

    Article  CAS  PubMed  Google Scholar 

  35. Roggo, C. & van der Meer, J. R. Miniaturized and integrated whole cell living bacterial sensors in field applicable autonomous devices. Curr. Opin. Biotechnol. 45, 24–33 (2017).

    Article  CAS  PubMed  Google Scholar 

  36. Kim, B. C. & Gu, M. B. A bioluminescent sensor for high throughput toxicity classification. Biosens. Bioelectron. 18, 1015–1021 (2003).

    Article  CAS  PubMed  Google Scholar 

  37. Cevenini, L., Calabretta, M. M., Tarantino, G., Michelini, E. & Roda, A. Smartphone-interfaced 3D printed toxicity biosensor integrating bioluminescent ‘sentinel cells’. Sens. Actuators, B Chem. 225, 249–257 (2016).

    Article  CAS  Google Scholar 

  38. Mimee, M. et al. An ingestible bacterial-electronic system to monitor gastrointestinal health. Science 360, 915–918 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Pedersen, M. W. et al. Ancient and modern environmental DNA. Philos. Trans. R. Soc. B. Biol. Sci. 370, 20130383 (2015).

    Article  Google Scholar 

  40. Sheth, R. U. & Wang, H. H. DNA-based memory devices for recording cellular events. Nat. Rev. Genet. 19, 718–732 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Farzadfard, F., Gharaei, N., Citorik, R. J. & Lu, T. K. Efficient retroelement-mediated DNA writing in bacteria. Preprint at bioRxiv https://doi.org/10.1101/2020.02.21.958983 (2020).

  42. Weiss, R. & Knight, T. F. Jr. in Cellular Computing (ed. Amos, M.) 120–121 (Oxford Univ. Press, 2004); https://doi.org/10.1093/oso/9780195155396.003.0012

  43. Chen, M. T. & Weiss, R. Artificial cell-cell communication in yeast Saccharomyces cerevisiae using signaling elements from Arabidopsis thaliana. Nat. Biotechnol. 23, 1551–1555 (2005).

    Article  CAS  PubMed  Google Scholar 

  44. Järup, L. & Åkesson, A. Current status of cadmium as an environmental health problem. Toxicol. Appl. Pharmacol. 238, 201–208 (2009).

    Article  PubMed  Google Scholar 

  45. Brocklehurst, K. R. et al. ZntR is a Zn(II)-responsive MerR-like transcriptional regulator of zntA in Escherichia coli. Mol. Microbiol. 31, 893–902 (1999).

    Article  CAS  PubMed  Google Scholar 

  46. Knierim, C., Greenblatt, C. L., Agarwal, S. & Greiner, A. Blocked bacteria escape by ATRP grafting of a PMMA shell on PVA microparticles. Macromol. Biosci. 14, 537–545 (2014).

    Article  CAS  PubMed  Google Scholar 

  47. Belkin, S. et al. Remote detection of buried landmines using a bacterial sensor. Nat. Biotechnol. 35, 308–310 (2017).

    Article  CAS  PubMed  Google Scholar 

  48. De Las Heras, A., Carreño, C. A. & De Lorenzo, V. Stable implantation of orthogonal sensor circuits in Gram-negative bacteria for environmental release. Environ. Microbiol. 10, 3305–3316 (2008).

    Article  Google Scholar 

  49. Tomović, N. S., Trifković, K. T., Rakin, M. P., Rakin, M. B. & Bugarski, B. M. Influence of compression speed and deformation percentage on mechanical properties of calcium alginate particles. Chem. Ind. Chem. Eng. Q. 21, 411–417 (2015).

    Article  Google Scholar 

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Acknowledgements

We thank F. Farzadfard for proving the SCRIBE strains and M. Mimee for providing the heme sensing strain. We thank S. Lin, N. Roquet, R. Citorik and S. Lemire for useful discussions. T.K.L. is grateful for funding received from the National Institutes of Health (NIH) New Innovator Award (no. 1DP2OD008435), NIH National Centers for Systems Biology (no. 1P50GM098792), the US Office of Naval Research (no. N00014-13-1-0424) and the Defense Advanced Research Projects Agency (no. HR0011-15-C-0091). X.Z. is grateful for funding received from the NIH (no. 1R01HL153857-01), the National Science Foundation (no. EFMA-1935291) and the US Army Research Office through the Institute for Soldier Nanotechnologies at MIT (no. W911NF-13-D-0001). C.F.-N. holds a Presidential Professorship at the University of Pennsylvania, is a recipient of the Langer Prize by the AIChE Foundation and acknowledges funding from the Institute for Diabetes, Obesity, and Metabolism, the Penn Mental Health AIDS Research Center of the University of Pennsylvania, the National Institute of General Medical Sciences of the NIH (no. R35GM138201), and the Defense Threat Reduction Agency (no. HDTRA11810041 and HDTRA1-21-1-0014). T.-C.T. gratefully acknowledge the support from The Abdul Latif Jameel Water and Food Systems Laboratory (J-WAFS) Graduate Student Fellowship.

Author information

Author notes

  1. Kevin Yehl

    Present address: Department of Chemistry and Biochemistry, Miami University, Oxford, OH, USA

  2. Cesar de la Fuente-Nunez

    Present address: Machine Biology Group, Departments of Psychiatry and Microbiology, Institute for Biomedical Informatics, Institute for Translational Medicine and Therapeutics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA

  3. Cesar de la Fuente-Nunez

    Present address: Departments of Bioengineering and Chemical and Biomolecular Engineering, School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, USA

  4. Cesar de la Fuente-Nunez

    Present address: Penn Institute for Computational Science, University of Pennsylvania, Philadelphia, PA, USA

  5. These authors contributed equally: Tzu-Chieh Tang, Eléonore Tham, Xinyue Liu.

Authors and Affiliations

  1. Synthetic Biology Group, Research Laboratory of Electronics, Massachusetts Institute of Technology, Cambridge, MA, USA

    Tzu-Chieh Tang, Eléonore Tham, Kevin Yehl & Timothy K. Lu

  2. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA

    Tzu-Chieh Tang, Kevin Yehl & Timothy K. Lu

  3. The Mediated Matter Group, Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA, USA

    Tzu-Chieh Tang

  4. Department of Materials Science and Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA

    Eléonore Tham

  5. Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA

    Xinyue Liu, Hyunwoo Yuk & Xuanhe Zhao

  6. Wyss Institute for Biologically Inspired Engineering, Boston, MA, USA

    Alexis J. Rovner

  7. Department of Genetics, Harvard Medical School, Harvard University, Boston, MA, USA

    Alexis J. Rovner

  8. Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT, USA

    Farren J. Isaacs

  9. Systems Biology Institute, Yale University, West Haven, CT, USA

    Farren J. Isaacs

  10. Department of Biomedical Engineering, Yale University, New Haven, CT, USA

    Farren J. Isaacs

  11. Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA

    Xuanhe Zhao

  12. Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, MA, USA

    Timothy K. Lu

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Contributions

T.-C.T., E.T., X.L., X.Z. and T.K.L. conceived and designed the research. T.-C.T., E.T., X.L. and H.Y. performed encapsulation and mechanical testing experiments. T.-C.T. and E.T. performed genetic circuit experiments. T.-C.T., E.T. and A.J.R. performed GRO experiments. T.-C.T. and E.T. performed river water experiments. T.-C.T., E.T., X.L., K.Y., A.J.R., C.F.-N., F.J.I., X.Z. and T.K.L. analyzed the data and wrote the manuscript.

Corresponding authors

Correspondence to Tzu-Chieh Tang, Xuanhe Zhao or Timothy K. Lu.

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Competing interests

T.-C.T., E.T., X.L., H.Y., X.Z. and T.K.L. have filed a patent application based on the hydrogel encapsulation technologies with the US Patent and Trademark Office. T.K.L. is a cofounder of Senti Biosciences, Synlogic, Engine Biosciences, Tango Therapeutics, Corvium, BiomX, Eligo Biosciences, Bota.Bio, Avendesora and NE47Bio. T.K.L. also holds financial interests in nest.bio, Armata, IndieBio, MedicusTek, Quark Biosciences, Personal Genomics, Thryve, Lexent Bio, MitoLab, Vulcan, Serotiny, Avendesora and Pulmobiotics.

Additional information

Peer review information Nature Chemical Biology thanks Keekyoung Kim, Minglin Ma and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Responses of encapsulated bacterial cells to external stimuli.

(a) Left: Schematic of GFP expression under the control of an aTc-inducible promoter. Center: Flow cytometry analysis of GFP expression in liquid culture and in hydrogel beads. Samples prepared in triplicate, data represent the mean ±1 s.d. based on analyses of 30000 events. The percentage data were calculated by dividing the numbers of GFP ON cells with the total cell counts. The fold-change data were derived from the mean of fluorescence. Right: Confocal microscopy images of beads encapsulating the aTc-sensing E. coli strain with and without 200 ng/mL aTc. (b) Left: A heme sensing strain which sense heme and generate bioluminescence as an output. The heme released from blood is transported into the cell by ChuA. Middle: Cells retrieved from beads showed a significant increase in luciferase activity. Right: The resulting bioluminescence can be detected with high sensitivity from intact beads. Samples prepared in triplicate, data represent the mean ±1 s.d.

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Supplementary Information

Supplementary Figs. 1–18 and Tables 1 and 2.

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Source Data Fig. 3

Statistical source data and loading curves.

Source Data Fig. 4

Statistical source data.

Source Data Fig. 5

Statistical source data.

Source Data Extended Data Fig. 1

Statistical source data.

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Tang, TC., Tham, E., Liu, X. et al. Hydrogel-based biocontainment of bacteria for continuous sensing and computation. Nat Chem Biol 17, 724–731 (2021). https://doi.org/10.1038/s41589-021-00779-6

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