Post‐proteomic identification of a novel phage‐encoded streptodornase, Sda1, in invasive M1T1 Streptococcus pyogenes

Abstract

SummaryThe M1T1 strain remains the most frequently isolated strain from group A streptococcal (GAS) infection cases worldwide. We previously reported that M1T1 differs from the fully sequenced M1 SF370 strain. To better understand the reason for the persistence and increased virulence of M1T1, we analysed its secreted proteome and identified two virulence proteins that are not present in the sequenced M1 SF370 strain: streptococcal pyrogenic exotoxin A (SpeA) and a streptodornase D (SdaD) homologue. In the present study, we determined the nucleotide sequence of the M1T1 streptodornase and found that its deduced amino acid sequence is highly similar to other streptococcal streptodornases, and is most closely related to the SdaD of GAS strain M49. M1T1 Sda shares two highly conserved domains with several DNases and putative DNases in streptococci; however, it possesses a unique C-terminal amino acid sequence. Thus, we named the protein Sda1, and we detected the presence of the sda1 gene in 16 M1T1 clinical isolates. The cloned and expressed Sda1 degrades both streptococcal and mammalian DNA at physiological pH. Amino acid similarity analyses of known GAS deoxyribonucleases suggest that Sda1 may be a chimeric protein created through recombination events. Moreover, a natural mutation that resulted in longer Sda1 and SdaD as compared to other GAS DNases was found to confer increased activity on the protein. Analysis of the sequences flanking sda1 determined that it is carried by a prophage or a prophage-like element inserted in the tRNA-Ser gene of M1T1 GAS. Ongoing studies in our laboratory aim to determine the contribution of Sda1 to the virulence of this globally disseminated M1T1 strain.

Rise and Persistence of Global M1T1 Clone ofStreptococcus pyogenes

“…After assembling these distinct sequences, we identifi ed 2 novel prophages that were introduced into the M1T1 global strain (19). One prophage (SPhinX) carries the speA2 gene, which encodes the potent superantigen SpeA; the other (PhiRamid) carries the sda1 gene, which encodes the most potent streptococcal nuclease identifi ed thus far (19,20). The introduction of these phages into the M1T1 clonal strain was later confi rmed by the complete genome sequence of a clinical M1T1 isolate, MGAS5005 (21).…”

Systems Biology Approach to Identifying Host Interactive Pathways Modulating the Severity of Streptococcal Sepsis

“…Another example of GAS interfering with PMN killing mechanisms is Sda1, a potent DNase produced by M1T1 (GAS strain under study). Sda1 dissolves neutrophil extracellular traps (NETs) allowing the bacteria to escape the entrapment and killing by PMN (Aziz et al, 2004b;Buchanan et al, 2006;Walker et al, 2007). Studies analyzing the role of macrophages in modulating differential susceptibility to GAS, using transcriptome analyses of GAS infected macrophages; found that GAS stimulates an "unusual" macrophage response.…”

“…Another example of GAS interfering with neutrophil killing mechanisms is Sda1, a potent DNase produced by M1T1 (GAS strain under study). Sda1 dissolves neutrophil extracellular traps (NETs), which are DNA filaments produced by neutrophils and contain histones and antimicrobial peptides, allowing the bacteria to escape the entrapment and on mechanism of killing by neutrophil (Aziz et al, 2004b;Buchanan et al, 2006;Walker et al, 2007).…”

“…Groups of 5-26 mice from a total of 30 BXD lines and their parental strains (B6 and D2) were injected via tail vein with 2  1.5 x 10 7 CFU/mouse of a hypervirulent form of the clinical isolate GAS 5448 (M1T1) Aziz et al, 2004a;Aziz et al, 2004b) as detailed previously (Aziz et al, 2007). Mortality and weight loss were recorded every 8 h for the ensuing 6 days.…”

Genome-wide analysis of Streptococcus pyogenes associated with pharyngitis and skin infections. (c2015)

“…High sequence homology was found between the genes in questions and their counterparts on bacteriophages, namely on the S. pyogenes phages 315.3, 315.6, and phage 9. Even though such genes were traditionally encoded on bacteriophages (Aziz et al, 2004), the uniformity of their location on the bacterial genomes suggests that they have become fixed remnants of phage integration events over time. Additionally, the Sda gene, a 981 bp fragment encoding the streptodornase B nuclease, was detected in two thirds of the isolates.…”