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The Parallel Cultivation Platform – Case study 1

Our Parallel Cultivation platform allows the cultivation and investigation of a wide range of microbial communities in extremely well-defined conditions. In this article, the use of this platform is exemplified by a case study on the storage of polymers by micro-organisms under different temperatures.

By the Parallel Cultivation Platform / July 28, 2021

KEY MESSAGES

The advent of techniques to quantify microbial genomes, has enabled researchers to characterize the composition of microbial communities in a wide range of ecosystems. What is lacking still, is insight in the functional performance of these communities. Linking community structure to function, would improve the understanding of microbial ecosystems and its applications. It is true that well-controlled bioreactors allow the investigation of both microbial community structure and function, but sampling and analysis of communities can interfere with bioreactor operation. Additionally, comparison of datasets from different studies with varying parameters, is limited by differences in study design. Clearly, there’s a need for a platform such as the Parallel Cultivation Platform, in which structure and function can be studied under different conditions in parallel.

Case study: Linking microbial community dynamics and functionality

To exemplify the use of the Parallel Cultivation Platform, we decribe a recently published study at – a.o. – the Delft University of Technology (TUD, citation below). In this work, we investigated the effect of temperature on microbial competition in parallel-operated reactors. The mixed microbial community in these bioreactors were pulse-fed with a single nutrient. As has been shown in previous studies, microbes compete for the limiting nutrient with at least two strategies: The “growers” directly convert the nutrient into biomass, whereas the “hoarders” store the nutrient as internal storage compounds (polymers) and convert them to biomass at a later timepoint.

PHA-storing bacteria

Hoarders have a competitive advantage over growers, as production of storage polymers is simpler than biomass production and consumes less energy. The enrichtment of storage polymer-producing bacteria is of great industrial potential, especially in the field of resource recovery from waste streams. What kind of storage polymer is formed, depends on the nutrient. Waste streams generally contain a lot of volatile fatty acids, which are converted by bacteria to polyhydroxyalkanoates (PHA). In turn, PHA can be used in the production of bioplastics and biochemicals. The picture below shows microbes that have stored PHA. Polyhydroxybutyrate (PHB) is the most commonly produced type of PHA by different bacteria.
Microscopy picture of microbial cells with PHA inclusion bodies, able to store up to 90% of the dry weight as PHA. Figure adopted from Stouten et al. (2019).

Experimental set-up

To study the effect of temperature on PHB-storing bacteria, eight parallel bioreactors were pulse-fed with acetate and operated in the temperature range of 20°C to 40°C. The sequencing batch bioreactors were operated for over 100 days and were online monitored with state-of-the-art equipment. Variables measured included O2 and CO2-off gas, base and acid dose, bioreactor temperature, dissolved oxygen and pH. Online data was used to reconstruct functional development of the enrichment cultures, as was verified by offline measurements.
Online and offline data of a single bioreactor cycle, where the online data could be used to a large extent to describe the functional behaviour of the enrichment culture. The gradual enrichment of hoarding biomass in the bioreactors could be monitored over the 200+ consecutive cycles. Figure adopted from Stouten et al. (2019).

Data processing and analyis

By utilizing all online data, it was possible to gain insights in the functional development of the enrichment cultures over time. A new data processing pipeline was developed to improve data handling and data interpretation by converting online measurements to real time microbial activity. For each operational cycle of each bioreactor the microbial functionality was captured as a mathematic depiction of the microbial activity. Three characteristic indicators were derived that showed strong correlations with microbial community structure development and offline-verified measurements, as reflected in the figure below for one bioreactor system.
Detailed overview of the experimental setup of reactors in the Parallel Cultivation Platform at TUD. This is shown to exemplify the conceptual set-up of the different reactor types included in UNLOCK. Figure adopted from Stouten et al. (2019).

Major benefits for this case study

Major benefits of the UNLOCK Parallel Cultivation Platform for this research included the ability to operate eight bioreactors in parallel from the same inoculum at high resolution. The online data processing tools, combined with the community structure analysis by 16s rRNA gene amplicon sequencing allowed for a novel experimental design and aided in the understanding of competitive strategies in microbial communities.

Interesting links

Major benefits of the UNLOCK Parallel Cultivation Platform

Read more about our Parallel Cultivation Platform.

Described study: Temperature as competitive strategy determining factor in pulse-fed aerobic bioreactors. The ISME Journal (2019) by G.R. Stouten, C. Hogendoorn, S. Douwenga, E.S. Kilias, G. Muyzer and R. Kleerebezem.

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