Orals

Nowadays, industrial fermentations rely almost entirely on the use of planktonic cells. However, biofilms (the most common form of bacterial growth in nature), offer several advantages to be exploited in modern fermentation processes. Bacteria in biofilms are more tolerant to several stresses than free cells, including toxic chemicals and shear stress. Furthermore, the adhesion of cells to surfaces can be exploited to operate a continuous fermentation process without excessive loss of biomass, thereby facilitating downstream processing. A programmable switch between planktonic and biofilm lifestyle is desirable to harness the advantages of both lifestyles. On this premise, we constructed a genetic gene circuit for biofilm formation in the platform strains Pseudomonas putida and Pseudomonas taiwanensis. Both P. putida and P. taiwanensis are robust, non-pathogenic soil bacteria and promising chassis for biotechnology as they can thrive under harsh operating conditions, displaying high tolerance towards several chemicals and can metabolize a broad range of substrates. These characteristics make them ideal for the production of a wide spectrum of chemicals. The synthetic circuit initiates biofilm formation upon detection of substrate or intermediate metabolites of the desired biotransformation, thus no additional inducer is needed. The circuit also allows for the propagation of cells in planktonic state prior employment in the bioreactor, which facilitates handling and speed up expansion of the culture. The design proposed herein employs a feedback-resistant diguanylate cyclase (DGC) from Caulobacter crescentus, which increases the concentration of DGC and therefore triggers biofilm formation. The resulting engineered strains were utilized for the biotransformation and degradation of chemicals (cyclohexanol) in continuous cultivation systems. This approach led to a ~300-fold increase in biofilm formation in microtiter plates, and was successfully used in diverse fermentation systems displaying increased catalytic efficiency.

How to cite: Volke, D. C., Heuschkel, I., Bühler, K., and Nikel, P. I.: Synthetic gene circuits for programmable Pseudomonas catalytic biofilms, biofilms 9 conference, 29 Sep–1 Oct 2020, biofilms9-128, https://doi.org/10.5194/biofilms9-128, 2020.

Erosion resistance is one of the advantages bacteria gain by producing biofilms. While it is undesirable for us humans when biofilms grow on medical devices or industrial pipelines, biofilms with a high erosion resistance can be advantageous for biotechnological applications. Here, we demonstrate how the erosion resistance of B. subtilis NCIB 3610 biofilms can be enhanced by integrating foreign (bio)polymers such as γ-polyglutamate (PGA), alginate and polyethylene glycol (PEG) into the matrix during biofilm growth.

Artificial enrichment of the NCIB 3610 biofilms with these biopolymers causes a significant increase in the erosion resistance by slightly changing the surface topography: A decreased cavity depth on the surface results in an alteration in the mode of surface superhydrophobicity, and we obtain a state that is located somewhere between rose-petal like and lotus-like wetting resistance. Surprisingly, the viscoelastic and microscopic penetration properties of the biofilms are not affected by the artificial incorporation of (bio)polymers. As we obtained similar results with all the biopolymers tested (which differ in terms of charge and molecular weight), this indicates that a variety of different (bio)polymers can be employed for a similar purpose.

The method introduced here may present a promising strategy for engineering beneficial biofilms such, that they become more stable towards shear forces caused by flowing water but, at the same time, remain permeable to nutrients or other molecules.

How to cite: Hayta, E. N. and Lieleg, O.: Enhanced erosion resistance of biopolymer-enriched B. subtilis NCIB 3610 biofilms, biofilms 9 conference, 29 Sep–1 Oct 2020, biofilms9-56, https://doi.org/10.5194/biofilms9-56, 2020.

Biofilms are considered as hotspots for the transfer of antibiotic resistance genes (ARGs), but very few studies have investigated the fate of ARGs (e.g. proliferation or elimination) in situ given different microbial spatial self-organization (SSO). SSO refers to a pervasive process during biofilm formation when microbes arrange themselves non-randomly across surfaces. So far the causes of SSO have been uncovered in a sense, however, the consequences of SSO were largely overlooked. Here, I hypothesize that the magnitude of inter-species intermixing, as one fundamental character of SSO, will determine the fate of ARG-carrying conjugative plasmid in both absence and presence of antibiotic selection. I evaluated this by performing range expansion experiments on agar plates to develop an artificial biofilm using a synthetic microbial community consisting of two isogenic Pseudomonas Stutzeri A1501 who are facultative denitrifiers in anaerobic condition. By knocking out different functional genes responsible for different steps of denitrification I am able to modify the metabolic interactions between these two strains from competing (without trophic interaction) to cross-feeding (with trophic interaction), which will further result in different magnitude of inter-species intermixing. Competing group has lower magnitude due to demixing of two, while cross-feeding group has higher magnitude due to mixing. I observed that in the absence of antibiotic selection plasmid experienced faster pace of elimination in competing group than cross-feeding group, whereas in the presence of antibiotic selection plasmid proliferated more efficiently in cross-feeding group than competing group. These results suggest that SSO is a determining factor of the fate of ARGs in biofilms, which provides a novel perspective of better understanding ARGs-related pressing problems facing our society.

How to cite: Ma, Y. and Johnson, D.: The effect of synthetic microbial spatial self-organization on the fate of antibiotic resistance genes, biofilms 9 conference, 29 Sep–1 Oct 2020, biofilms9-72, https://doi.org/10.5194/biofilms9-72, 2020.