Oral Presentation BACPATH 2019

Scarlet fever-causing Streptococcus pyogenes require streptolysin O, host cytosolic glutathione and prophage exotoxins for nasopharyngeal infection (#3)

Stephan Brouwer 1 , Timothy C Barnett 1 , Diane Ly 2 , Katherine J Kasper 3 , David MP De Oliveira 1 , Tania Rivera-Hernandez 1 , Amanda J Cork 1 , Johanna Richter 1 , Gordon Dougan 4 5 , Victor Nizet 6 , Kwok-Yung Yuen 7 , John K McCormick 3 8 , Martina L Sanderson-Smith 2 , Mark R Davies 1 9 , Mark J Walker 1
  1. Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, QLD, Australia
  2. Ilawarra Health and Medical Research Institute and School of Biological Sciences, University of Wollongong, Wollongong, New South Wales, Australia
  3. Department of Microbiology and Immunology and the Centre for Human Immunology, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada
  4. The Wellcome Trust Sanger Institute, Hinxton, Cambridge, United Kingdom
  5. University of Cambridge, Cambridge, United Kingdom
  6. Department of Pediatrics, University of California San Diego, La Jolla, California, USA
  7. State Key Laboratory for Emerging Infectious Diseases, The University of Hong Kong, Hong Kong Special Administrative Region, Hong Kong , China
  8. Lawson Health Research Institute, London, Ontario, Canada
  9. Department of Microbiology and Immunology at the Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Melbourne, VIC, Australia

The re-emergence of scarlet fever since 2011 poses a new threat to global public health. A detailed understanding of the processes controlling the upsurge in scarlet fever cases is therefore instrumental in the fight against this disease. The capacity of North Asian serotype M12 (emm12) Streptococcus pyogenes (Group A Streptococcus, GAS) to cause scarlet fever has been linked to the presence of antibiotic resistance elements and novel prophages, including prophage ΦHKU.vir encoding the secreted superantigens SSA, SpeC and the DNase Spd1. However, robust empirical evidence defining the contribution of prophage-encoded exotoxins to the pathogenesis of scarlet fever is lacking. Here we sought to investigate the regulation of these exotoxins and what role they play in GAS nasopharyngeal infection using defined genetic knockouts in a humanized mouse model for pharyngeal infection. A small molecule screen identified biothiols, such as glutathione (GSH), as a factor specifically enhancing secretion and activity of the exotoxin SSA. Our data reveal SSA as the first thiol-activated superantigen to be reported. We show that GSH is released from host cellular stores through the action of the pore-forming toxin streptolysin O, providing a mechanistic framework for how extracellular GAS gains access to highly abundant GSH in vivo. This work also provides evidence that DNase Spd1 is required for optimal growth in human blood and confers resistance to neutrophil killing. To examine how these exotoxins contribute to the fitness of GAS, we generated isogenic single and triple knockout mutants in the ssa, speC and spd1 genes. Loss of all three exotoxins significantly reduced nasopharyngeal colonization in a mouse infection model, whereas single mutants did not, suggesting a synergistic relationship between these exotoxins. Taken together, these findings offer a new paradigm for GAS pathogenesis and support the hypothesis that acquisition of ΦHKU.vir-encoded exotoxins has played a key role in the selection and expansion of scarlet fever lineages in North Asia.