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Biofilms – complex microbial communities and their molecular tricks

Microorganisms can be found in drinking water processing systems, catheter cannulae and implants – habitats where they can cause serious problems for human health. It is not an easy task to get rid of the bacteria as they form bacterial communities that constitute an excellent protection against UV radiation, disinfectants and antibiotics. A group of researchers led by Dr. Thomas Schwartz at the Karlsruhe Institute of Technology (KIT) is investigating the composition of biofilms and how the microbes react to external attacks. Micro- and molecular biology methods are used to gather information about the well-organised and complex bacterial communities. How do the bacterial colonisers react to stress? Can the colonisation of cannulae, implants and drinking water systems be prevented by interfering with bacterial signalling pathways, for example?

The formation of a biofilm consists of several stages: It begins with the attachment of free-floating bacteria, unicellular fungi or algae to a surface. These early colonisers then produce a layer of polysaccharides, proteins and nucleic acids (known as extracellular polymeric substances, EPS). The EPS initially form a two-dimensional layer that is difficult to remove mechanically. As the biofilm matures, the bacterial or fungal colony becomes three-dimensional, new species colonise the area, subpopulations develop and take on a broad range of different tasks. This leads to aerobic and anaerobic areas containing different bacterial and fungal species with different biochemical behaviour. “A mature biofilm is not an arbitrary system consisting of individual bacteria,” said Dr. Thomas Schwartz from the Institute of Functional Interfaces at the Karlsruhe Institute of Technology (KIT). “The cells exchange signalling molecules and react to each other”, said Schwartz referring to how the bacteria communicate on a chemical level, thus enabling them to coordinate gene expression according to the local density of the bacterial population. This phenomenon is referred to as quorum sensing. A biofilm is very effective in protecting the bacteria against external influences. Even though the outer layers of biofilm can be destroyed with disinfectants, antibiotics and UV, other parts of the biofilm will survive and start spreading again.

Schematic showing the five stages of biofilm formation: attachment of molecules, attachment of bacteria, microcolonies, biofilm maturation and EPS synthesis, maturation and release<br />
Schematic of biofilm development on a surface © Dr. Thomas Schwartz

Weapons to counteract stress

Schwartz and his group of researchers in the Department of Microbiology of Natural and Technical Interfaces are working with partners from academia and industry to investigate the processes of biofilms that are mainly formed by bacteria. The researchers’ goal is to gain a detailed understanding of the development and organisation of such complex ecosystems. They hope that by identifying relevant molecular mechanisms, they will be able to come up with approaches to control the development of clinically and industrially relevant biofilms. Each investigation starts with the elucidation of biofilm composition. Using genetic fingerprinting, Schwartz’s team first establish a taxonomic profile of a biofilm community in order to identify the individual species in a particular biofilm. The next step involves the use of molecular biology methods to identify genes that are expressed during the colonisation of surfaces or during biofilm development as well as the associated molecular signalling processes. This step is followed by the search for unknown bacterial genes and for genetic programmes that are switched on to counteract stress, all of which are mechanisms used by the microbes to defend themselves against antibiotics, UV radiation, etc.

Microscopic image of a biofilm stained with a fluorescence dye © Dr. Thomas Schwartz

Results from one particular research project are a good example of how Schwartz and his team are working towards their goal. Drinking water is generally disinfected with UV emitters. UV radiation breaks up the structure of bacterial DNA, thereby inactivating the bacteria contained in drinking water. However, some of the bacteria that colonise the tubes and tanks of a drinking water purification system can evade these attacks by activating mechanisms that are able to repair DNA damage. It is not very long before the bacteria are able to proliferate again, thus causing the contamination of drinking water to the extent that it becomes unfit for human consumption. “For example, projects which we have carried out in cooperation with drinking water and waste management partners have shown that some UV-irradiated bacteria switch on the gene recA,” said Schwartz. “This gene activates proteins that attach to damaged DNA stretches in order to repair them.” This enables some of the irradiated bacteria to survive and form new bacterial communities. In order to remove bacterial contamination, care needs to be taken to completely and permanently destroy all undesired bacteria.

Tough opponents and new surfaces

RecA is one of numerous bacterial genes that control the molecular response to stress. It is interesting to note that recA also has another function: the induction of recA expression by stress such as strong UV radiation or contact with antibiotics also leads to an increase in the rate at which individual bacteria in a biofilm exchange DNA with each other. This process, known as horizontal gene transfer, is based on the ability of microorganisms to take up free DNA from their environment and incorporate it into their own genome. This enables the bacteria to exchange the capacity to become resistant to antibiotics, thus making bacteria quite tough opponents: What does not kill them, makes them stronger. “We need to understand the molecular mechanisms that contribute to preserving the biofilm. This is the only way to effectively prevent bacteria from colonising implants or drinking water purification systems in the future,” said Schwarz.

The search for key genetic regulators of bacterial stress responses is a potential approach towards eliminating undesired bacterial colonisers. Another approach is trying to find a way to prevent the formation of biofilm altogether. In future this approach might become feasible through the development of new materials whose surface properties prevent, or at least delay, the bacterial colonisation of surfaces. Schwartz and his team are focusing on such approaches, working together with industrial partners or colleagues from the KIT. In a project carried out in cooperation with BASF, the researchers are investigating certain substances produced by fungi. These substances, mainly proteins, have the potential to modify the chemical properties of material surfaces or to destabilise the structure of established biofilms, thereby reducing or preventing the risk of bacterial biofilm formation.

It is still not known whether the aforementioned methods are suitable for effectively preventing the colonisation of implants and drinking water systems with bacteria. However, the better researchers understand the complex processes that lead to the development of a biofilm, the better their ability to direct their strategies to eradicating undesired bacterial communities.

Further information:

Dr. Thomas Schwartz
Group: Microbiology of Natural and Technical Interfaces
Institute of Functional Interfaces (IFG)
Karlsruhe Institute of Technology (KIT)
Tel.: +49-721/608-2-6802
E-mail: thomas.schwartz(at)kit.edu

Website address: https://www.biooekonomie-bw.de/en/articles/news/biofilms-complex-microbial-communities-and-their-molecular-tricks