Plasmids are circular molecules of DNA that can replicate autonomously in bacterial cells. The current genetic engineering technology is based on plasmids that contain an origin of replication and a resistance marker and can be introduced into bacterial recipient strain by transformation.

The PI of this project invented a new type of plasmid vectors (pFOS) that do not contain antibiotic resistance genes as selection markers but can be selected by a diet enriched with scFOS, short chain fructooligosaccharides. The scFOS are a group of linear fructose oligomers already available as prebiotics on the market. These sugars can escape digestion in the human upper intestine, reach the colon, and are not metabolized by most of the species belonging to the Enterobacteriaceae family. Metabolizing these sugars, a non-pathogenic Escherichia coli strain carrying the pFOS vector, is thought to have a selective advantage against other enterobacteria, transiently colonizing the gut. In addition, the pFOS vector is self-conjugative, thus has the capacity to be transferred from one bacterial strain to another bacterial recipient cell. The final pFOS technology is thought to be used in vivo to bring genetic information within a complex bacterial population (i.e., the intestinal microbiota), vehiculating genetic information to resident, autochthonous bacteria colonizing the gut of humans and animals. This project aims to construct a second-generation of pFOS plasmid variants with improved conjugation abilities and a wider bacterial host range and to measure by fermentation of cultivated microbiota the conditions of selection and stability of the constructs. Such in vitro study is fundamental to describe the microbiota response to scFOS and to select the most appropriate plasmid to become a good genetic vector for future applications of microbiome genetic engineering. The application of these vectors may help to target antimicrobial-resistant bacteria colonizing the gut of animals and humans.