Transmission of Antimicrobial Resistance by GEne Transfer within Bacterial Biofilms – TARGET-biofilms

Bacterial communities are composed of a variety of species, which have the ability to exchange genetic material through horizontal gene transfer (HGT), thus accelerating their diversification, adaptation and genetic evolution. The major gene transfer mechanism is bacterial conjugation, where DNA is transferred from a donor to a recipient cell by direct contact. The importance of conjugation has first become obvious after the Second World War, with the global spread of multidrug resistance. Nowadays, analysis of drug-resistant commensal, environmental and pathogen strains has revealed a vast collection of conjugative plasmids carrying one or multiple resistance genes to most, if not all classes of antibiotics currently used in clinical treatments. Conjugation-mediated drug-resistance dissemination is consequently a major obstacle to successful treatment of infections and is now recognized as one of the biggest threats in public health.

In natural and clinical environments, bacterial species predominantly live in spatially structured communities called biofilms, in which a self-produced extracellular matrix holds cells together. Biofilms offer protection against potentially toxic compounds such as antimicrobial drugs, thus intrinsically limiting the efficiency of conventional antimicrobial therapy. Biofilms also protect the bacterial community against bacteriophage infection, rendering phage-based therapeutic approaches unsuitable. Importantly, it has been reported that the biofilm mode of life facilitates horizontal genes transfer by conjugation. Therefore, biofilms not only shelter bacteria against external hazards, but could also offer a niche that facilitates the dissemination of drug-resistance determinants. This possibility highlights the need to invest research effort into the accumulation of basic knowledge about the dynamics of gene transfer by DNA conjugation within bacterial biofilm communities.

The central goal of TARGET-Biofilms research proposal is to explore the fundamental and practical aspect of DNA conjugation within bacterial biofilms. The first objective is to provide a comprehensive understanding of the mechanisms, extent, and impact of bacterial conjugation within biofilms. To do so, we will use a genetic system to monitor intra- and interspecies cell-to-cell DNA transfer in live-cells, in combination with an experimental set-up allowing direct imaging of bacterial biofilms in 3D at cellular and sub-cellular resolution. We will address the influence of the biofilm structure and strain composition on the efficiency of DNA transfer and investigate the dissemination of drug-resistance within the community. The second objective is to test the possibility to combine conjugation and CRISPR systems to perform in situ biofilm manipulation, including attempts to disassemble biofilms by killing the resident cells or by suppressing genes required for biofilm formation and stability. Combining DNA transfer functions of conjugation systems with CRISPR/Cas9 genes designed to have a specific effect in targeted bacterial species is an unexplored, yet promising approach, which deserves high interest.
The research program will benefit from the consortium’s expertise in live-cell microscopy approaches dedicated to the study of DNA transfer and biofilms, as well as molecular microbiology. TARGET-Biofilms research program will provide outcomes beneficial to public health, life stock, crops, and the environment.