Beyond the emm-type: fine-tuning Group A Streptococcus typing with Enn and Mrp

Regarding the increased tendency to look at the M and M-like proteins as a functional trio, we set up a PCR-based typing of the enn and mrp genes in Group A Streptococcus (GAS). Based on a large mga regulon database, we have designed specific PCR primers for amplification and sequencing of enn and mrp genes. Primers were first validated in silico using a global database of 1,419 GAS genomes and then in vitro on 17 isolates covering all subclusters of Enn and Mrp proteins. Off-targets were also checked against other streptococci harboring M-like genes. We also described how to assign new Enn- and Mrp-types based on an up-to-date database available online. This new typing system will enhance the precision of strain characterization and help understand some differences between isolates sharing the same M-type.IMPORTANCEStreptococcus pyogenes, also known as Group A Streptococcus, is a bacterium that causes a wide range of diseases, from mild sore throats to life-threatening infections and long-term complications like rheumatic heart disease. It is responsible for over 500,000 deaths a year. For decades, scientists have focused on the surface M protein which helps it survive in human tissues. The large diversity of its N-terminal region has led to the emm-typing system of S. pyogenes. However, the two M-like proteins, Enn and Mrp, produced by the same region of the genome, have been largely overlooked until recently. These proteins are found in many strains around the world, and recent findings suggest they also play an important role in S. pyogenes virulence. In our study, we developed a new typing system for Enn and Mrp proteins that aims to better understand their behavior and improve the monitoring of clinical strains.