Periodic Reporting for period 1 - BACKSPiN (Bacteriocins and human commensal bacteria as a new strategy to inhibit the opportunistic human pathogen Streptococcus pneumoniae)
Período documentado: 2021-05-01 hasta 2023-04-30
When the Noble Prize-winning Alexandre Fleming discovered antibiotics, he already noticed that microorganisms developed resistance mechanisms to survive. Therefore, he anticipated that a misuse of antimicrobial compounds to treat infections will drive the selection of hyper-resistant strains and the resurgence of almost-eradicated infectious diseases. Nowadays, the problem is so critical that the World Health Organization foresees that superbugs will outcompete cancer and cardiovascular diseases to become the first cause of mortality on the planet in less than 30 years (horizon 2050). Recently, the international organization drew a list of 10 priority pathogens that includes Streptococcus pneumoniae, a bacterium notorious in pneumonia (major upper respiratory tract infections), endocarditis, meningitis and brain abscess. To replace or restore antibiotic action, we thus need to find alternative strategies. In this proposal, I aim to use bacteriocins, small antimicrobial peptides secreted by bacteria, to kill S. pneumoniae. They are currently underexploited for human need but feature many valuable characteristics (e.g. efficiency, evolvability, specific spectrum, cheap/easy production, high sequence diversity, stability) complementary to antibiotics. I will test a collection of hundreds of bacteriocins and, according to their mode of action, will rationally assemble “overwhelming” bacteriocin cocktails to prevent emergence of resistance. In parallel, a tantalizing idea would be to exploit the beneficial bacteria of our microbiota and mobilize their bacteriocins to treat local infections. So, I will perform ex vivo infection of human epithelia with S. pneumoniae and test how bacteriocin-induced S. salivarius, a commensal bacterium of our gut, influences it. Besides generating valuable fundamental insight into the S. pneumoniae resistance mechanisms and infection cycle, the results of this project will also pave the way to fight against other notorious pathogens.
In order to identify new bacteriocin-based therapies, I selected a commensal bacterium, Streptococcus salivarius, and tested its probiotic properties on S. pneumoniae in conditions activating the production of bacteriocins. In parallel, I screened a library of chemically synthesized bacteriocins and tested their inhibitory properties on S. pneumoniae viability. Moreover, I used a pooled, high-throughput screening approach called CRISPRi to monitor the effect of potent bacteriocins on a library of S. pneumoniae knock-down mutants. CRISPRi (Clustered Regularly Interspaced Short Palindromic Repeats Interference) relies on genome-wide gene depletions and enables to highlight genetic interactions in specific condition (i.e. bacteriocin treatments) at the genome scale.
In this project, I have successfully tested a combination of bacteriocin producer strains or a combination of synthetic bacteriocins. I identified several bacteriocins or cocktails of bacteriocins able to inhibit S. pneumoniae growth at workable concentrations. In order to propose efficient infection therapies on a long run, I deeply investigated the mode of action of these bacteriocins and the mechanism of resistance of S. pneumoniae. I discovered that a protease played a generalist role in the protection against peptide-based antimicrobials.
In this project, I have successfully tested a combination of bacteriocin producer strains or a combination of synthetic bacteriocins. I identified several bacteriocins or cocktails of bacteriocins able to inhibit S. pneumoniae growth at workable concentrations. In order to propose efficient infection therapies on a long run, I deeply investigated the mode of action of these bacteriocins and the mechanism of resistance of S. pneumoniae. I discovered that a protease played a generalist role in the protection against peptide-based antimicrobials.
The different conditions I tested to inhibit S. pneumoniae were very promising. I show that 50% of the S. salivarius strains were able to kill this pathogen while many synthetic bacteriocins (with 6 highly potent ones) were effective on plate and in liquid. Moreover, some bacteriocin modules prevent the emergence of resistance. This project will lead to a patent to protect at least 2 bacteriocins that could be used to treat infectious caused by pathogenic bacteria in clinical contexts.