CRISPR Inhibition of Prophage Acquisition in Streptococcus pyogenes

Takashi Nozawa,Ichiro Nakagawa,Nayuta Furukawa,Bijaya Haobam,Chihiro Aikawa,Ken Kurokawa,Takayasu Watanabe,Fumito Maruyama,Eliane Namie Miyaji

doi:10.1371/journal.pone.0019543

Abstract

Streptococcus pyogenes, one of the major human pathogens, is a unique species since it has acquired diverse strain-specific virulence properties mainly through the acquisition of streptococcal prophages. In addition, S. pyogenes possesses clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems that can restrict horizontal gene transfer (HGT) including phage insertion. Therefore, it was of interest to examine the relationship between CRISPR and acquisition of prophages in S. pyogenes. Although two distinct CRISPR loci were found in S. pyogenes, some strains lacked CRISPR and these strains possess significantly more prophages than CRISPR harboring strains. We also found that the number of spacers of S. pyogenes CRISPR was less than for other streptococci. The demonstrated spacer contents, however, suggested that the CRISPR appear to limit phage insertions. In addition, we found a significant inverse correlation between the number of spacers and prophages in S. pyogenes. It was therefore suggested that S. pyogenes CRISPR have permitted phage insertion by lacking its own spacers. Interestingly, in two closely related S. pyogenes strains (SSI-1 and MGAS315), CRISPR activity appeared to be impaired following the insertion of phage genomes into the repeat sequences. Detailed analysis of this prophage insertion site suggested that MGAS315 is the ancestral strain of SSI-1. As a result of analysis of 35 additional streptococcal genomes, it was suggested that the influences of the CRISPR on the phage insertion vary among species even within the same genus. Our results suggested that limitations in CRISPR content could explain the characteristic acquisition of prophages and might contribute to strain-specific pathogenesis in S. pyogenes.

Highlights

During evolution, bacteria acquired new traits primarily by horizontal gene transfer (HGT) as a key driving force for expressing novel pathogenic properties, new colonization niches as well as metabolic adaptations [1,2,3,4,5]
The CRISPR1 terminal repeat-like sequence of MGAS10750 is located between hemN and lepA, those of MGAS315 and SSI-1 are distant by 39.5 kb from the CRISPR1 locus of the other strains (Fig. S1A, B)
S. pyogenes strains are well-known to induce a variety of diseases and the sequences of 13 strains suggested that the prophage regions contain variable virulence genes and confer pathogenic capacities [31]

Summary

Introduction

Bacteria acquired new traits primarily by horizontal gene transfer (HGT) as a key driving force for expressing novel pathogenic properties, new colonization niches as well as metabolic adaptations [1,2,3,4,5]. Conjugation, transduction and transformation are the major mechanisms for HGT. Of the three HGT mechanisms, transduction involving bacteriophage-mediated DNA transfer often provides the profound alteration in host bacterial genomes. This process can convert a non-pathogenic strain into a pathogenic variety through prophage-encoded toxins, surface alterations, or increasing resistance to human immunity [7]. Prophage insertion into the host genome often inactivates or alters the host genes [7,8]

Methods

Results

Conclusion