PT  - JOURNAL ARTICLE
AU  - Nathan Tague
AU  - Haonan Lin
AU  - Jean-Baptiste Lugagne
AU  - Deeya Burman
AU  - Wilson W. Wong
AU  - Ji-Xin Cheng
AU  - Mary J. Dunlop
TI  - Single-cell chemical imaging of engineered strains reveals heterogeneity in fatty acid production
AID  - 10.1101/2021.07.26.453865
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.07.26.453865
4099  - http://biorxiv.org/content/early/2021/07/26/2021.07.26.453865.short
4100  - http://biorxiv.org/content/early/2021/07/26/2021.07.26.453865.full
AB  - Engineered microbes can produce valuable chemicals, however production strains often require extensive optimization before they can be used at industrial scales. To quantify production, metabolic engineers typically employ mass spectrometry, which offers excellent chemical specificity. However, with this approach samples are derived from bulk cultures, obscuring potential cell-to-cell differences in production, and precluding selection methods that rely on measurements of a single cell’s phenotype, such as directed evolution. Here, we use stimulated Raman scattering (SRS) based chemical imaging to directly visualize free fatty acids in metabolically engineered Escherichia coli. We uncover substantial heterogeneity in fatty acid production among and within colonies. We then demonstrate longitudinal SRS imaging, allowing fatty acids to be linked with the cells that generated them. Further, using the hyperspectral images, we develop an approach for compositional analysis that determines the chain length and unsaturation of the fatty acids in living cells. Our results demonstrate that cell-to-cell heterogeneity in production of metabolic targets can be substantial and indicate that controlling this to eliminate low producers represents a critical area for future optimization. Further, because SRS imaging is rapid, chemically specific, and can be acquired longitudinally, it is a promising method for metabolic engineers, allowing for direct imaging of biosynthesis at the single-cell level.Competing Interest StatementThe authors have declared no competing interest.