Broad-range lytic bacteriophages that kill Staphylococcus aureus local field strains

Abstract

Staphylococcus aureus is a very successful opportunistic pathogen capable of causing a variety of diseases ranging from mild skin infections to life-threatening sepsis, meningitis and pneumonia. Its ability to display numerous virulence mechanisms matches its skill to display resistance to several antibiotics, including β-lactams, underscoring the fact that new anti-S. aureus drugs are urgently required. In this scenario, the utilization of lytic bacteriophages that kill bacteria in a genus -or even species- specific way, has become an attractive field of study. In this report, we describe the isolation, characterization and sequencing of phages capable of killing S. aureus including methicillin resistant (MRSA) and multi-drug resistant S. aureus local strains from environmental, animal and human origin. Genome sequencing and bio-informatics analysis showed the absence of genes encoding virulence factors, toxins or antibiotic resistance determinants. Of note, there was a high similarity between our set of phages to others described in the literature such as phage K. Considering that reported phages were obtained in different continents, it seems plausible that there is a commonality of genetic features that are needed for optimum, broad host range anti-staphylococcal activity of these related phages. Importantly, the high activity and broad host range of one of our phages underscores its promising value to control the presence of S. aureus in fomites, industry and hospital environments and eventually on animal and human skin. The development of a cocktail of the reported lytic phages active against S. aureus-currently under way- is thus, a sensible strategy against this pathogen.

Fig 1. Transmission electron micrographs.

Images correspond to phages, A) vB_Sau_S24, B) vB_Sau_CG and C) vB_Sau_Clo6.

Fig 2. Genome organization of the phage vB_Sau_S24.

ORFs functions are shown in different colors. The 72 putative promoters were represented by an arrow and 34 putative rho-independent transcriptional terminators by stem loops.

Fig 3. Genome organization of the phage vB_Sau_CG.

ORFs functions are shown in different colors. The 70 putative promoters were represented by an arrow and 31 putative rho-independent transcriptional terminators by stem loops.

Fig 4. Genome organization of the phage vB_Sau_Clo6.

ORFs functions are shown in different colors. The 70 putative promoters were represented by an arrow and 31 putative rho-independent transcriptional terminators by stem loops.

Fig 5. SplitsTree networks analysis of the Myoviridae family bacteriophages.

Thirty genomes available in database were aligned by Gegenees and the network was built with SplitsTree4 by the Neighbor-Net method. Five phylogenetic groups (clusters) were identified. vB_Sau_S24 and vB_Sau_Clo6 clustered together forming a separate group.

Fig 6. CCT map comparing the genome of phage vB_Sau_S24 to other S. aureus myobacteriophages.

The most external ring in the graph corresponds to vB_Sau_S24 used as a reference genome. The next 32 rings correspond to BLASTN alignment of the each genome analyzed, the color correspond to the percent of sequence similarity (see below panel).

Fig 7. CCT map comparing the genome of phages vB_Sau_CG to other S. aureus myobacteriophages.

The most external ring in the graph corresponds to vB_Sau_CG used as a reference genome. The next 32 rings correspond to BLASTN alignment of the each genome analyzed, the color correspond to the percent of sequence similarity (see below panel).

Fig 8. CCT map comparing the genome of phage vB_Sau_Clo6 to other S. aureus myobacteriophages.

The most external ring in the graph corresponds to vB_Sau_Clo6 used as a reference genome. The next 32 rings correspond to BLASTN alignment of the each genome analyzed, the color correspond to the percent of sequence similarity (see below panel).