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Acoustic reporter genes for noninvasive imaging of microorganisms in mammalian hosts

Bourdeau, Raymond W. and Lee-Gosselin, Audrey and Lakshmanan, Anupama and Farhadi, Arash and Kumar, Sripriya Ravindra and Nety, Suchita P. and Shapiro, Mikhail G. (2018) Acoustic reporter genes for noninvasive imaging of microorganisms in mammalian hosts. Nature, 553 . pp. 86-90. ISSN 0028-0836. http://resolver.caltech.edu/CaltechAUTHORS:20171102-124711465

[img] Image (JPEG) (Extended Data Figure 1 : Sequence homology of GvpA/B) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 2 : Ultrasound contrast from buoyancy-enriched cells) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 3 : Time course of acoustic reporter gene contrast after induction) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 4 : Acoustic reporter gene expression and ultrasound imaging does not affect cell viability) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 5 : Multiplexed imaging of genetically engineered reporter variants) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 6 : Anatomical ultrasound images of acoustic bacteria in the gastrointestinal tract) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 7 : Ultrasound imaging of ARG-expressing cells in the mouse colon) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 8 : Effect of arg1 and lux expression on ECN cell growth, viability and microcin release) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 9 : Ultrasound imaging of S. typhimurium in tumour xenografts) - Supplemental Material
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[img] Image (JPEG) (Extended Data Figure 10 : High-throughput screening of acoustic phenotypes) - Supplemental Material
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[img] PDF (Supplementary Notes 1–5, Plasmid Sequences for ARG1 and ARG2 constructs, Supplementary References and Supplementary Table 1) - Supplemental Material
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Abstract

The mammalian microbiome has many important roles in health and disease1,2, and genetic engineering is enabling the development of microbial therapeutics and diagnostics3,4,5,6,7. A key determinant of the activity of both natural and engineered microorganisms in vivo is their location within the host organism8,9. However, existing methods for imaging cellular location and function, primarily based on optical reporter genes, have limited deep tissue performance owing to light scattering or require radioactive tracers10,11,12. Here we introduce acoustic reporter genes, which are genetic constructs that allow bacterial gene expression to be visualized in vivo using ultrasound, a widely available inexpensive technique with deep tissue penetration and high spatial resolution13,14,15. These constructs are based on gas vesicles, a unique class of gas-filled protein nanostructures that are expressed primarily in water-dwelling photosynthetic organisms as a means to regulate buoyancy16,17. Heterologous expression of engineered gene clusters encoding gas vesicles allows Escherichia coli and Salmonella typhimurium to be imaged noninvasively at volumetric densities below 0.01% with a resolution of less than 100 μm. We demonstrate the imaging of engineered cells in vivo in proof-of-concept models of gastrointestinal and tumour localization, and develop acoustically distinct reporters that enable multiplexed imaging of cellular populations. This technology equips microbial cells with a means to be visualized deep inside mammalian hosts, facilitating the study of the mammalian microbiome and the development of diagnostic and therapeutic cellular agents.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://dx.doi.org/10.1038/nature25021DOIArticle
https://www.nature.com/articles/nature25021PublisherArticle
ORCID:
AuthorORCID
Bourdeau, Raymond W.0000-0003-2202-1980
Lakshmanan, Anupama0000-0002-6702-837X
Shapiro, Mikhail G.0000-0002-0291-4215
Additional Information:© 2018 Macmillan Publishers Limited, part of Springer Nature. Received 21 December 2016; accepted 9 November 2017. We thank F. S. Foster, D. Maresca, A. Mukherjee, M. Din, T. Danino, J. Willmann and S. K. Mazmanian for discussions, and A. McDowall for assistance with electron microscopy. This research was supported by the National Institutes of Health grant R01-EB018975, the Canadian Institute of Health Research grant MOP 136842 and the Pew Scholarship in the Biomedical Sciences. A.L. is supported by the NSF graduate research fellowship (award 1144469) and the Biotechnology Leaders Program. A.F. is supported by the NSERC graduate fellowship. S.P.N. was supported by the Caltech Summer Undergraduate Research Fellowship. Research in the Shapiro laboratory is also supported by the Heritage Medical Research Institute, the Burroughs Wellcome Career Award at the Scientific Interface and the David and Lucile Packard Fellowship for Science and Engineering. Author Contributions: R.W.B. and M.G.S. conceived and designed the study. R.W.B., A.L., A.L.-G., A.F. and S.P.N. prepared genetic constructs in E. coli. R.W.B., A.L., A.L.-G., S.P.N. and A.F. conducted in vitro ultrasound experiments. A.L.-G. and R.W.B. performed in vivo ultrasound experiments. A.L., A.F. and A.L.-G. conducted metabolic burden experiments in Nissle 1917 cells. R.W.B. and S.R.K. prepared genetic constructs in S. typhimurium. R.W.B. and A.L. obtained TEM images. R.W.B., A.L.-G. and M.G.S. analysed ultrasound data. R.W.B. and M.G.S. wrote the manuscript with input from all authors. M.G.S. supervised the research. Code availability: MATLAB code is available from the corresponding author upon reasonable request. Data and code availability: arg1 and arg2 plasmid sequences are included in Supplementary Information, and plasmids will be available from Addgene. All other materials are available from the corresponding author upon reasonable request. The authors declare no competing financial interests.
Group:Heritage Medical Research Institute
Funders:
Funding AgencyGrant Number
NIHR01-EB018975
Canadian Institutes of Health Research (CIHR)MOP 136842
Pew Charitable TrustUNSPECIFIED
NSF Graduate Research FellowshipDGE-1144469
Biotechnology Leaders ProgramUNSPECIFIED
Natural Sciences and Engineering Research Council of Canada (NSERC)UNSPECIFIED
Caltech Summer Undergraduate Research Fellowship (SURF)UNSPECIFIED
Heritage Medical Research InstituteUNSPECIFIED
Burroughs Wellcome FundUNSPECIFIED
David and Lucile Packard FoundationUNSPECIFIED
Record Number:CaltechAUTHORS:20171102-124711465
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20171102-124711465
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ID Code:82898
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:02 Jan 2018 22:22
Last Modified:03 Jan 2018 18:24

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