The Zeigler Report

Summary: We interviewed Nick Zeigler, a professional working with Brewdog in the beer industry. He gave us many an insight on how our project could be applied in industry. There is a need for robust yet dynamic mixed cultures in industries ranging from waste water treatment and biofuel production to brewery. The concept of designing bio-feedback loops for establishing a self regulating system in a flow-through reactor was discussed, and two examples for the application of such a system were given. The first example was about the management of carbon metabolism in biofuel production, and the second was about the accumulation of propionic acid as a byproduct during the breakdown of cellulosic materials in water treatment. These are discussed in further detail below. He also discussed the requirement for rapid feedback from the reactor, in order to monitor its status, and the possibility of having static populations of “buffer” strains. These buffer strains would “wait in the wings” of the bioreactor and degrade harmful byproducts as they occurred, increasing the robustness of the community. Ideally they would also be reporting the levels of these byproducts.

Waste water treatment

One major issue that waste water treatment plants need to face is the variation in the feedstock. Municipal waste varies greatly through the year, often in unpredictable ways. A spike in acidity or fat content, for example, may destabilise the composition of the bioreactor irreversibly, by killing off one or more of the populations. Robust organisms that will survive a range of chemical environments are therefore required, but so is a method for controlling the natural fluctuations of the population composition, so that it does not override and permanently destabilise the community within the bioreactor. In the cellulose degradation example, a common byproduct that becomes prevalent is propionic acid. This compound is toxic for most cells except syntrophic propionate oxidising bacteria (SPOB). These organisms degrade propionic acid and can be used to buffer the reactor medium. When there is a spike in propionate, SPOBs no longer buffer the medium and instead overtake it, and subsequently die as the propionate is depleted. Current methods for assaying the composition of a bioreactor can take as long as a day, which is not fast enough to prevent the collapse of the microbial community if the composition has unbalanced. Therefore there is a need for a reporting device, that can inform us about the activity of the organisms in the bioreactor, such as the activity of SPOBs. This could readily be measured by detecting a secondary metabolite of SPOBs. Ideally, that would be coupled with a mechanism for adjusting the system accordingly. For example, if the main producer is making lots of propionate, causing it to accumulate, that would cause an explosion in SPOBs. If a third strain, which would act as a reporter strain, detects the increased activity of SPOBs and reports it to the user, then adjustments can be made to the feedstock. Furthermore, the third strain could be appropriately picked to use the SPOBs’ byproduct, converting it to something that can be used by the main producers. This would enhance their growth and prevent them from being overrun by SPOBs. Such a system would in effect be autoregulatory, thus minimizing the need for manual interference and bioreactor down time. An alternative mechanism for preventing the collapse of the microbial community, would be to synthetically impose a limit on the growth of SPOBs. By enforcing a method of population control, SPOBs can continue to remove propionate without the risk of them taking over the bioreactor. Using this method, we impose a hierarchy on the system, where the main producer always takes precedent in the bioreactor, over the “buffer” strains.

Biofuel production

The second issue which arose is the preferential metabolism of simple sugars (such as glucose) over more complex ones. A common bioreactor setup for biofuel production consists of a series of tanks, with a different set of metabolic transformations occurring in each tank. Having the first tank deal with the simple sugars, allows the more complex ones to be dealt with by the microorganisms in the next tank. The organisms in the second tank are usually specialised to degrade complex sugars, but will still preferentially metabolise simple sugar if fed it. What happens if there is a spike in simple sugar influx? The cells in the first tank will not be able to metabolise all of it, leading to leakage of sugars to the next reactor. This will have an inhibitory effect on the metabolism of the more complex sugars (which is slow to recover), and can upset the reactions downstream. The suggested solutions were to modify the carbon metabolism of these cells using synthetic biology, most likely to improve recovery speed once all simple sugars have been metabolised.