Poster Presentation 20th Lancefield International Symposium on Streptococci and Streptococcal Diseases 2017

Functional Analysis of Streptococcal Virulence Factors (#161)

Kar Yan Soh 1 , Jacelyn Loh 1 2 , Chris Hall 1 , Thomas Proft 1 2
  1. School of Medical Science, The University of Auckland, Auckland, New Zealand
  2. Maurice Wilkins Center, The University of Auckland, Auckland, New Zealand

Background: Streptococcus pyogenes (Group A Streptococcus, GAS) causes a variety of diseases in humans ranging from pharyngitis and impetigo to more severe invasive diseases including cellulitis, necrotising fasciitis and toxic shock. Investigations of GAS pathomechanisms are hindered by the lack of suitable animal infection models. Objectives: To characterise the mechanisms of two important GAS virulence factors, Streptococcus pyogenes nuclease A (SpnA) and streptococcal 5’-nucleotidase A (S5nA), by investigating the orthologues SpnAi and S5nAi in the fish pathogen Streptococcus iniae using a zebrafish infection model. Results: SpnAi and S5nAi were cloned and expressed in E. coli. Biochemical analysis of purified rSpnAi and rS5nAi showed that both proteins have very similar reaction conditions compared to SpnA and S5nA. rSpnAi is able to digest linear double-stranded DNA and chromosomal DNA with highest activity at pH 6.5–7.5 and between 32°C–37°C in the presence of Ca2+ and Mg2+. rS5nAi is able to generate immunomodulatory molecules adenosine and macrophage toxic deoxyadenosine with highest activity at pH 7 and 42°C in the presence of Mg2+. A S. iniae spnAi gene deletion mutant has been generated by allelic replacement and is ready for analysis in a zebrafish infection model. Conclusion: SpnAi and S5nAi have been confirmed as true orthologues of the GAS proteins SpnA and S5nA, respectively. The study of these proteins in the zebrafish infection model will provide novel insights into the virulence mechanisms that might be correlated to the in-vivo functions of the orthologue GAS proteins in the human host.