Characterization of the genetic switch from phage ɸ13 important for Staphylococcus aureus colonization in humans.

Camilla S Kristensen,Hanne Ingmer,Mogens Kilstrup,Helena A K Leinweber,Leila Lo Leggio,Karin Hammer,Anders K Varming

doi:10.1002/mbo3.1245

Camilla S Kristensen, Hanne Ingmer + Show 5 more

Open Access

https://doi.org/10.1002/mbo3.1245

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Abstract

Temperate phages are bacterial viruses that after infection either reside integrated into a bacterial genome as prophages forming lysogens or multiply in a lytic lifecycle. The decision between lifestyles is determined by a switch involving a phage‐encoded repressor, CI, and a promoter region from which lytic and lysogenic genes are divergently transcribed. Here, we investigate the switch of phage ɸ13 from the human pathogen Staphylococcus aureus. ɸ13 encodes several virulence factors and is prevalent in S. aureus strains colonizing humans. We show that the ɸ13 switch harbors a cI gene, a predicted mor (modulator of repression) gene, and three high‐affinity operator sites binding CI. To quantify the decision between lytic and lysogenic lifestyle, we introduced reporter plasmids that carry the 1.3 kb switch region from ɸ13 with the lytic promoter fused to lacZ into S. aureus and Bacillus subtilis. Analysis of β‐galactosidase expression indicated that decision frequency is independent of host factors. The white “lysogenic” phenotype, which relies on the expression of cI, could be switched to a stable blue “lytic” phenotype by DNA damaging agents. We have characterized lifestyle decisions of phage ɸ13, and our approach may be applied to other temperate phages encoding virulence factors in S. aureus.

Highlights

Staphylococcus aureus is a Gram-positive, opportunistic human pathogen causing millions of infections worldwide each year, ranging from skin and soft tissue infections, food poisoning, endocarditis, and respiratory tract infections to bacteremia, amongst many others (Liu, 2009; Lowy, 1998; Tong et al, 2015)
E. coli and Bacillus subtilis were grown in lysogeny broth (LB) or LB agar, while S. aureus was grown in tryptic soy broth (TSB) or agar (TSA)
3.1 | Topology of the ɸ13 switch region and detection of identical switches in members of the Sa3int and the PVL encoding phages residues from the peptidase domain as well as the autocatalytic site from λ are conserved in ɸ13, suggesting that the CI repressor from ɸ13, in contrast to the CI repressor from TP901-1 (Madsen et al, 1999), could be capable of autocleavage

Summary

| INTRODUCTION

Staphylococcus aureus is a Gram-positive, opportunistic human pathogen causing millions of infections worldwide each year, ranging from skin and soft tissue infections, food poisoning, endocarditis, and respiratory tract infections to bacteremia, amongst many others (Liu, 2009; Lowy, 1998; Tong et al, 2015). As a member of the staphylococcal Sa3int phage family, the bacteriophage ɸ13 shares features with the other members These features include the Sa3 type integrase and the presence of the sak, chp, and scn virulence genes (Goerke et al, 2009; Xia & Wolz, 2014), but an analysis of the genetic switches within the group has never been reported. By constructing plasmids containing the ɸ13 genetic switch with lacZ reporter fusions to the lytic promoter (switch plasmids) and analysis of the frequency of Lac phenotypes in transformants of the natural and a heterologous host, we have shown that a functional mor gene is required for “decision switching” and that this process is independent of staphylococcal host factors. The results may aid in the understanding of how some Sa3int prophages are disseminated in society and suggest ways to limit S. aureus colonization in humans

| EXPERIMENTAL PROCEDURES

| RESULTS AND DISCUSSION

| CONCLUSIONS