Engineering bacterial hosts cells for robust growth at high external osmolarities
Lead Research Organisation:
University of Edinburgh
Abstract
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People |
ORCID iD |
Publications
Buda R
(2016)
Dynamics of Escherichia coli's passive response to a sudden decrease in external osmolarity.
in Proceedings of the National Academy of Sciences of the United States of America
Stevenson K
(2016)
General calibration of microbial growth in microplate readers.
in Scientific reports
Swain PS
(2016)
Inferring time derivatives including cell growth rates using Gaussian processes.
in Nature communications
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| BB/M015777/1 | 30/06/2015 | 29/06/2019 | |||
| 1602789 | Studentship | BB/M015777/1 | 30/06/2015 | 29/06/2019 |
| Description | We have identified the limits of optical density measurements of bacterial suspensions in microplate readers that arise from the complex relationship of size, refractive index and number of bacteria. From this study we have developed a method for the calibration of cell number in these readers that takes into account these limits allowing for continued use of microplate readers as a high throughput data collection method. We have also developed an automated cell counting protocol to improve throughput of calibration as well as collect data on cell size distributions. Using a microplatereader and our calibration methodology we have devised a protocol for the rapid scanning of osmolarities, medium and solvent concentrations to identify optimal conditions for growth and production. In our studies we have used exposure to ethanol as an example of a toxic product of an industrial bioreactor and have looked to identify conditions that improve the resistance of our E.coli. We have so far concluded that the introduction of high osmolarity in combination with exposure to ethanol causes a reduction in the ethanol tolerance of the bacteria, likely due to the significant simultaneous stress imposed by both conditions. In addition, we have however identified that a reduction in temperature significantly improves solvent tolerance, with a 2% increase in resistance by reducing the incubation temperature 10C. |
| Exploitation Route | We hope that the calibration technique for microplate readers we have developed will be used as a standard protocol for future researchers to help provide accurate measurements of cell number when using optical density measurements. We also intend to continue with our research into the use of osmoregulation systems for the improvement of bacteria for industrial production of compounds, including synthetic gene constructs for over expression of various components of the osmoregulation pathways. In addition we are now beginning to measure the diffusion within the cytoplasm of the bacteria at different osmolarties in order to better understand the reduction of growth rate that occurs with the intention of finding novel ways to mitigate this. In addition, reduction of growth temperature may be useful in certain processes for improving solvent tolerances of the bacteria, however, the reduced growth rate that results may offset that advantage and thus some further research is needed. |
| Sectors | Manufacturing, including Industrial Biotechology |
| URL | https://www.nature.com/articles/srep38828 |
| Description | A physiological approach to understanding osmotically induced growth modulation |
| Amount | £175,636 (GBP) |
| Funding ID | RPG-2019-187 |
| Organisation | The Leverhulme Trust |
| Sector | Charity/Non Profit |
| Country | United Kingdom |
| Start | 03/2020 |
| End | 02/2023 |
| Description | EPSRC IAA |
| Amount | £55,360 (GBP) |
| Funding ID | EPSRC IAA PIII065 |
| Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 09/2019 |
| End | 12/2020 |
| Description | HFSP program grant |
| Amount | $1,000,000 (USD) |
| Funding ID | RGP0041/2015 |
| Organisation | Human Frontier Science Program (HFSP) |
| Sector | Charity/Non Profit |
| Country | France |
| Start | 05/2015 |
| End | 12/2018 |
| Description | Industrial CASE Account - University of Edinburgh 2019 |
| Amount | £257,700 (GBP) |
| Funding ID | EP/T517501/1 |
| Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 08/2019 |
| End | 09/2024 |
| Description | Collaboration with Chr Hansen |
| Organisation | Chr. Hansen A/S |
| Country | Denmark |
| Sector | Private |
| PI Contribution | In this collaboration, we are looking at the effects of osmolarity changes on the survival of prokaryotic bacteria. |
| Collaborator Contribution | This is confidential information. |
| Impact | Currently, there has been one company visit by student Mark Zurbruegg for the purpose of data collection, we are currently analysing the data. |
| Start Year | 2019 |
| Description | Collaboration with Matt Scott at University of Waterloo |
| Organisation | University of Waterloo |
| Country | Canada |
| Sector | Academic/University |
| PI Contribution | This partnership was lead by our team in Edinburgh where we performed microscopy to measure the rates of diffusion in the E.coli cytoplasm and contributed to the development of a model describing E.coli growth at high osmolarity, building on the collaborators previous work. |
| Collaborator Contribution | Our partners provided training and expertise in experimental protocols for the analysis of proteome fractions within bacterial cells and useful guidance in developing the model. |
| Impact | No output yet |
| Start Year | 2018 |
| Description | Collaboration with Peking University, group of Prof. Fan Bai |
| Organisation | Peking University |
| Department | Biodynamics Optical Imaging Center |
| Country | China |
| Sector | Academic/University |
| PI Contribution | This partnership was lead by our team in Edinburgh where the team at Peking University contributed to the mathematical modeling of bacterial response to osmotic downshocks. The collaboration is ongoing. The two three undergraduate students who helped with the worked secured funding for PhD positions (two) and a Msc scholarship (one). |
| Collaborator Contribution | This partnership was lead by our team in Edinburgh where the team at Peking University contributed to the mathematical modeling of bacterial response to osmotic downshocks. The collaboration is ongoing. The two three undergraduate students who helped with the worked secured funding for PhD positions (two) and a Msc scholarship (one). |
| Impact | Buda R*, Liu Y*, Yang J*, Hegde S*, Stevenson K, Bai F** and Pilizota T**. Dynamics of Escherichia coli's passive response to a sudden decreases in external osmolarity. PNAS September 2016, doi:10.1073/pnas.1522185113 This collaboration is interdisciplinary, it is work that falls under biological physics. |
| Start Year | 2016 |
| Title | Cell counter |
| Description | The software automatically counts numbers of cells in a given sequence of images. |
| Type Of Technology | Software |
| Year Produced | 2018 |
| Impact | The software will likely be a part of a spin-out company and a device in development. We are currently looking into protecting it. |
| Title | Fitderiv |
| Description | The software is written to provide a tool for estimating growth rates from optical density data and uses that language (although it can process any other type of data too). It has been described in more detail in the following publication Swain P S**, Stevenson K, Leary A, Montano-Gutierrez L F, Clark I B N, Vogel J and Pilizota T. Inferring time-derivatives, including cell growth rates, using Gaussian processes. Nature Communications 2016;7:13766 and made openly available at http://swainlab.bio.ed.ac.uk/software/fitderiv/ |
| Type Of Technology | Software |
| Year Produced | 2016 |
| Open Source License? | Yes |
| Impact | The software is currently used by several research groups and expected to be adopted wider (publication date is December 2016) |
| URL | http://swainlab.bio.ed.ac.uk/software/fitderiv/ |