Antibiotic resistance in the important opportunistic human pathogen Streptococcus pneumoniae is on the rise. This is particularly problematic in the case of the β-
lactam antibiotic amoxicillin, which is the first-line
therapy. It is therefore crucial to uncover targets that would kill or resensitize
amoxicillin-resistant pneumococci. To do so, we developed a genome-wide, single-cell based, gene silencing screen using CRISPR interference called sCRilecs-seq (subsets of CRISPR interference libraries extracted by fluorescence activated cell sorting coupled to next generation sequencing). Since
amoxicillin affects growth and division, sCRilecs-seq was used to identify targets that are responsible for maintaining proper cell size. Our screen revealed that downregulation of the
mevalonate pathway leads to extensive cell elongation. Further investigation into this phenotype indicates that it is caused by a reduced availability of cell wall precursors at the site of cell wall synthesis due to a limitation in the production of
undecaprenyl phosphate (Und-P), the
lipid carrier that is responsible for transporting these precursors across the cell membrane. The data suggest that, whereas
peptidoglycan synthesis continues even with reduced Und-P levels, cell constriction is specifically halted. We successfully exploited this knowledge to create a combination treatment strategy where the FDA-approved
drug clomiphene, an inhibitor of Und-P synthesis, is paired up with
amoxicillin. Our results show that
clomiphene potentiates the antimicrobial activity of
amoxicillin and that combination
therapy resensitizes
amoxicillin-resistant S. pneumoniae. These findings could provide a starting point to develop a
solution for the increasing amount of hard-to-treat
amoxicillin-resistant
pneumococcal infections.