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Measuring the adhesion capacity between two bacteria using an AFM
Researchers at LAAS-CNRS and TBI have succeeded in characterizing, using an atomic force microscope (AFM), the role of protein filaments (pili) on the cell surface during the interaction between two bacteria. These results are published in the journal ACS Applied Materials & Interfaces.
Pili are protein filaments a few microns in length, present on the wall of some bacteria. They contribute to interactions between bacterial cells and play a role in the formation of biofilms, whether on biotic (mucous membranes, intestinal cells) or abiotic (polystyrene, stainless steel) surfaces. A team made up of researchers from our ELiA - Engineering in Life Science Applications team and the Toulouse biotechnology institute (TBI, CNRS/INRAE/INSA Toulouse)1, has succeeded in precisely measuring the adhesion forces between two bacteria linked to the presence of these pili, using an instrument derived from the atomic force microscope (AFM).
TBI researchers were studying the adhesion mechanisms of lactic acid bacteria, while those at LAAS-CNRS had the expertise in the use of the AFM microscope in biology. Together, they worked with a model bacterium - Lactococcus lactis - one of the most widely used lactic acid bacteria in the dairy industry, particularly in cheese and fermented dairy product manufacturing processes. To study the influence of pili on the adhesion capacity between two bacteria, they used three laboratory strains, which produce pili (pili+), overproduce pili (pili++), or weak pili (pili-).
Les mesures de l'interaction entre les bactéries ont été effectuées à l'aide d'une technique de spectroscopie de force cellulaire (Single-cell force spectroscopy, SCFS). Elle consiste à remplacer la pointe d'un microscope AFM par une boule de silice d'environ 5 microns de diamètre, sur laquelle une bactérie individuelle est fixée par collage. Ce dispositif a permis de mesurer sa force d'interaction avec les bactéries isolées sur une lame de microscope.
The experiments showed the existence of strong adhesion forces (150 to 452 pN) between bacteria for the strains expressing pili on their surface (pili+ and pili++), while these forces remained below 100 pN for the pili-weak strain (pili-). Similarly, the adhesion work, - the amount of energy required to separate two bacteria - exceeds 50.10-18 J for the pili+ and pili++ strains, while it is less than 10.10-18 J for the pili- strain. These results confirm the role of pili-pili interactions in the adhesion mechanism between these cells, an elementary building block in the structuring of a biofilm. They also show that pili interact with other proteins present on the surface of bacteria.
These partners have already moved on to the next phase of the study, which aims to highlight, beyond the interaction between two individual bacteria, the role of pili in the structuring and mechanical properties of a biofilm. A new publication is in preparation.
Interactions between two bacterial pili measured by force spectroscopy. The AFM lever is functionalized by a single bacterium and interacts with another bacterium immobilized on the surface. On the left, cells without pili do not interact with each other, resulting in flat force curves, while on the right, bacteria expressing pili interact with each other in several modes, characterized by the strength, distance and shape of the interactions.
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1 in collaboration with the MICALIS Institute (INRAe/AgroParisTech/University of Paris-Saclay)
Analysis of Homotypic Interactions of Lactococcus lactis Pili Using Single-Cell Force Spectroscopy,
I. Dramé, C. Formosa-Dague, C. Lafforgue, M-P. Chapot-Chartier, J-C Piard, M.Castelain, and E. Dague,
ACS Applied Materials & Interfaces (Avril 2020)
Etienne Dague, LAAS-CNRS, firstname.lastname@example.org
Mickael Castelain, TBI, email@example.com