Christian Lesterlin

Horizontal DNA transfer and the spread of drug resistance:

Understanding how a commensal bacterium becomes drug resistant through horizontal gene is a central issues in Microbiology. One way to address these questions is to use a combination of molecular genetics and microscopy imaging in live cells, which has proven to be a powerful strategy to gain insight into biological processes. Recent high-resolution imaging techniques, in particular Structured-Illumination Microscopy (SIM), enable us to observe the real-time dynamics of proteins or DNA sequences with unprecedented resolution. Live cell imaging technic is combined with specifically developed sample preparation and microfluidics apparatus that allow observation of cells growing under physiological conditions over several generations. Hence, fundamental mechanisms that previously have only been characterised through genetic or biochemical studies can now be investigated in the context of the living cell.

We then study the acquisition of DNA by bacterial conjugation at the cytological scale. Conjugation is the major process involved in the acquisition of exogenous DNA and associated new properties such as pathogenesis or multidrug resistance. Translating our molecular knowledge of conjugation mechanisms to an in vivo context is currently the biggest challenge in the field and is the main objective of our research. To tacle these questions, we have developed a genetic system that enables the microscopic visualization of each key steps of DNA conjugation in live cells, i.e., DNA transfer from donor to recipient cells, maintenance of the acquired DNA and expression of newly acquired gene in the transconjugant cells and consequent establishment of new properties, such as resistance to antibiotics. In particular, we study the dynamics of acquisition of a tetracycline-resistance-carrying plasmid in order to characterise the chronology of conversion of commensal bacterium into a drug resistant cell.