Streptococcus pyogenes sugar metabolism influences antimicrobial activity and niche-specific infection

Amanda C. Marple, The University of Western Ontario

Abstract

Streptococcus pyogenes is a human-adapted pathogen that causes a variety of infections including pharyngitis and skin infections, and although this bacterium produces many virulence and host colonization factors, how S. pyogenes competes with the host microbiota is not well understood. Here we detected antimicrobial activity produced from S. pyogenes MGAS8232 that was able to prevent the growth of Micrococcus luteus. This activity was produced when cells were grown in 5% CO₂ in M17 media supplemented with galactose; however, the addition of alternative sugars coupled with genome sequencing experiments revealed the antimicrobial phenotype was not related to classical bacteriocins. Ascorbic acid availability was also discovered to be important in the production of the antimicrobial phenotype. To further determine genes involved in the production of this activity, a transposon mutant library in S. pyogenes MGAS8232 was generated. The transposon screen identified the mannose phosphotransferase system (Man-PTS), a major sugar transporter in S. pyogenes, as important for the antimicrobial phenotype, which was also the secondary mutation identified in the bacteriocin-deficient (ΔspbJKMN) mutant. Additional loss-of-function transposon mutants were identified to also be involved in alternative sugar utilization. Sugar utilization profiles in all the Man-PTS mutants demonstrated that galactose, mannose, and N-acetylglucosamine utilization was impaired. In vitro RNA-seq experiments in high and low glucose concentrations further identified the Man-PTS as a glucose transporter; however, there was no transcriptional regulators or virulence factors affected with the loss of the Man-PTS. A clean deletion in the Man-PTS demonstrated defects in a mouse model of nasopharyngeal, but not in skin infection. We also developed a novel RNA isolation method of S. pyogenes-infected nasopharyngeal murine tissues. This elucidated alternative sugar import, virulence regulators and factors important during peak infection. This method also elucidated that the Man-PTS was important for the initial stages of nasal infection and not at 48 hours post-infection. Overall, the ability of S. pyogenes to utilize alternative sugars presented by glycans seems to play a role in the initial stages of acute infection and interactions with the endogenous microbial population existing in the nasopharynx.