The most crucial difference between a bacterial biofilm and free-floating (planktonic) single-cells living in liquid suspensions is the biofilm’s dramatically increased chemotolerance. This has been attributed to various factors; the protective extracellular matrix that can restrict antibiotics penetration, a different phenotype characterized by more tolerance-oriented mechanisms (e.g. efflux pumps) and the different metabolic states of the bacteria within the biofilm. In the core of the community reside bacterial cells with very low or no metabolic activity (persisters). They comprise about 1-5% of the bacterial population in both biofilms and planktonic stationary phase cultures and are known to be very tolerant to antimicrobials and can revive after treatment. They are the main cause of recalcitrance of chronic, biofilm related infections.
Biofilms are regarded as one of the greatest challenges of modern biomedicine and they are claimed responsible for a massive proportion (up to 80%) of the antibiotic-tolerant infections. They are involved in several chronic diseases such as lung pneumonia of cystic fibrosis patients, otitis media, chronic non-healing wounds and infection of medical implants and devices. Antimicrobial resistance and tolerance has become a recognized problem worldwide. There is a need for changes in the use of existing antimicrobials, but also a need for alternative therapies to be able to tackle this problem. Targeting virulence factors such as quorum sensing (QS), the system responsible for group behaviours in bacteria, is a strategy that reduces the selective pressure on the bacteria to survive and thus limits the development of resistance towards the treatment. Another strategy to consider is to use combination therapies and adjuvants to existing antibiotics. These alternative treatments are especially of need in therapies for biofilm infections, as there is still a very limited repertoire of molecules that can selectively act on mature biofilms. With the recognition of biofilm’s importance as the predominant state for bacterial living instead of the planktonic lifestyle, there has been a progressive shift towards a new era in drug discovery in which searching for antibiofilms is urged to occupy a more prominent place. Our group take part in that effort to find novel ways to study and fight biofilm infections.
We have established an assay platform for testing various compound libraries to evaluate their effect on viability, biomass and biofilm matrix (1). The platform was established using Staphylococcus aureus and S. epidermidis strains and has also been optimized for Pseudomonas aeruginosa, Burkholderia cenocepacia and Escherichia coli biofilm forming strains. This platform is microtiter-well plate-based and allows the evaluation of the activity of compounds on both the planktonic bacteria and the biofilms.
(Skogman et al., 2012)
Our research group also studies the biofilm colonization onto various surfaces, as well as assess the effectivity of antimicrobial biomaterials. We have developed and characterized in-depth two platforms that allows the study of biofilms onto different surfaces: the agar plate-based (3, 4) and the microtiter plate-based method. The bacterial biofilm can be quantify by viable cell density, biofilms matrix (carbohydrates and proteins) and biomass.
(Hiltunen et al., 2016)
We are also interested in different approaches to fight biofilms. Quorum sensing is the communication system of microorganisms that allows them to adopt community-based synchronised behaviours. As such, it plays an important role in biofilm formation. We have developed a microwell plate-based assay platform, using a reporter strain, Chromobacterium violaceum, to efficiently screen for quorum sensing inhibitors that can be further studied on relevant pathogens using the same quorum sensing system (5). We additionally seek to take the characterization of tolerant bacterial phenotypes further by applying proteomics approaches. By studying the changes in protein composition on bacterial cell surfaces or in the biofilm matrix during the processes leading to multi-drug tolerance, we can acquire insight e.g. for the development of bioactive materials and novel treatments for tackling bacterial persistence.
This consortium project is lead by Docent Adyary Fallarero with an overall funding of 640 000 euros. This translational project aims to advance early drug discovery research focused on the identification and characterisation of new Quorum Sensing Inhibitors, as novel anti-bioflm strategies. The main goal is to develop a set of new drug naturally-inspired candidates, acting as Quorum sensing inhibitors (QSI) of the LuxI/LuxR system, essential for Pseudomonas aeruginosa biofilm formation. The project will focus on:
- Mechanistic studies of QSI inhibitors and their ability to counteract biofilms
- Structural refinements of already identified QSI compounds
- In silico screens of QSI
- Combination studies of the most active leads with known antibiotics
This effort is a collaborative consortium involving also the groups of prof. Jari Yli-Kauhaluoma (medicinal chemistry) and Docent Henri Xhaard (computational discovery).
For more information, please contact: adyary.fallarero helsinki.fi
2017-2020, granted by the Academy of Finland
The consortium TWIN-A aims at advancing the transition into sustainable and competitive bioeconomy through creation of high value-added products in line with circular economy goals. TWIN-A project investigates low-value waste treatment processes as unprecedented sources of unique compounds against bacterial biofilm using advanced technologies.
2017-2020, granted by the European Union’s Horizon 2020 research and innovation programme
PRINT-AID is a European training network for development of personalized anti-infective medical devices combining printing technologies and antimicrobial functionality.
Within this network, our role has focused on: (i) the identification of repurposed investigational or approved drugs that inhibit bacterial colonisation or kill bacteria, the structural optimization and the characterization of the mode of action of the selected lead. (ii) The study of the antimicrobial functionality of the 3D-printed materials with the selected formulations or combinations and the development of assays for a better characterization of biofilm inhibitors intended to bio-fabrication of anti-infective bone implants. A new in vitro assay based on the co-culture of human cells and bacteria on the material of interest was developed with the aim to more closely resemble the conditions encountered by implants in vivo (6).
The University of Helsinki team consist of: Professor Jari Yli-Kauhaluoma (scientific coordinator), Dr. Pia Fyhrquist (project manager), Docent Adyary Fallarero (co-supervisor) and Docent Kirsi Savijoki (co-eupervisor)
1. Skogman, M.E.; Vuorela, P.M.; Fallarero, A. Combining biofilm matrix measurements with biomass and viability assays in susceptibility assessments of antimicrobials against Staphylococcus aureus biofilms. The Journal of antibiotics 2012, 65, 453-459, doi:10.1038/ja.2012.49.
2. Skogman, M.E.; Vuorela, P.M.; Fallarero, A. A Platform of Anti-biofilm Assays Suited to the Exploration of Natural Compound Libraries. J Vis Exp 2016, 10.3791/54829, doi:10.3791/54829.
3. Oja, T.; Blomqvist, B.; Buckingham-Meyer, K.; Goeres, D.; Vuorela, P.; Fallarero, A. Revisiting an agar-based plate method: what the static biofilm method can offer for biofilm research. Journal of microbiological methods 2014, 107, 157-160, doi:10.1016/j.mimet.2014.10.008.
4. Hiltunen, A.K.; Skogman, M.E.; Rosenqvist, K.; Juvonen, H.; Ihalainen, P.; Peltonen, J.; Juppo, A.; Fallarero, A. Bioactive glass combined with bisphosphonates provides protection against biofilms formed by the periodontal pathogen Aggregatibacter actinomycetemcomitans. International journal of pharmaceutics 2016, 501, 211-220, doi:10.1016/j.ijpharm.2016.02.006.
5. Skogman, M.E.; Kanerva, S.; Manner, S.; Vuorela, P.M.; Fallarero, A. Flavones as Quorum Sensing Inhibitors Identified by a Newly Optimized Screening Platform Using Chromobacterium violaceum as Reporter Bacteria. Molecules (Basel, Switzerland) 2016, 21, doi:10.3390/molecules21091211.
6. Reigada, I.; Perez-Tanoira, R.; Patel, J.Z.; Savijoki, K.; Yli-Kauhaluoma, J.; Kinnari, T.J.; Fallarero, A. Strategies to Prevent Biofilm Infections on Biomaterials: Effect of Novel Naturally-Derived Biofilm Inhibitors on a Competitive Colonization Model of Titanium by Staphylococcus aureus and SaOS-2 Cells. Microorganisms 2020, 8, doi:10.3390/microorganisms8030345.