Role of the pre-neck appendage protein (Dpo7) from phage vB_SepiS-philPLA7 as an anti-biofilm agent in Staphylococcal species

Staphylococcus epidermidis and Staphylococcus aureus are important causative agents of hospital-acquired infections and bacteremia, likely due to their ability to form biofilms. The production of a dense exopolysaccharide (EPS) matrix enclosing the cells slows the penetration of antibiotic down, resulting in therapy failure. The EPS depolymerase (Dpo7) derived from bacteriophage vB_SepiS-philPLA7, was overexpressed in Escherichia coli and characterized. A dose dependent but time independent response was observed after treatment of staphylococcal 24 h-biofilms with Dpo7. Maximum removal (>90%) of biofilm-attached cells was obtained with 0.15 mu M of Dpo7 in all polysaccharide producer strains but Dpo7 failed to eliminate polysaccharide-independent biofilm formed by S. aureus V329. Moreover, the pretreatment of polystyrene surfaces with Dpo7 reduced the biofilm biomass by 53-85% in the 67% of the tested strains. This study supports the use of phage-encoded EPS depolymerases to prevent and disperse staphylococcal biofilms, thereby making bacteria more susceptible to the action of antimicrobials

Characterization of novel bacteriophages for biocontrol of bacterial blight in leek caused by Pseudomonas syringae pv. porri

Pseudomonas syringae pv. porri, the causative agent of bacterial blight in leek (Allium porrum), is increasingly frequent causing problems in leek cultivation. Because of the current lack of control measures, novel bacteriophages were isolated to control this pathogen using phage therapy. Five novel phages were isolated from infected fields in Flanders (vB_PsyM_KIL1, vB_PsyM_KIL2, vB_PsyM_KIL3, vB_PsyM_KIL4, and vB_PsyM_KIL5), and were complemented with one selected host range mutant phage (vB_PsyM_KIL3b). Genome analysis of the phages revealed genome sizes between 90 and 94 kb and an average GC-content of 44.8%. Phylogenomic networking classified them into a novel clade, named the "KIL-like viruses," related to the Felixounalikevirus genus, together with phage phiPsa374 from P. syringae pv. actinidiae. In vitro characterization demonstrated the stability and lytic potential of these phages. Host range analysis confirmed heterogeneity within P. syringae pv. porri, leading to the development of a phage cocktail with a range that covers the entire set of 41 strains tested. Specific bio-assays demonstrated the in planta efficacy of phages vB_PsyM_KIL1, vB_PsyM_KIL2, vB_PsyM_KIL3, and vB_PsyM_KIL3b. In addition, two parallel field trial experiments on three locations using a phage cocktail of the six phages showed variable results. In one trial, symptom development was attenuated. These data suggest some potential for phage therapy in controlling bacterial blight of leek, pending optimization of formulation and application methods

Antibacterial phage ORFans of Pseudomonas aeruginosa phage LUZ24 reveal a novel MvaT inhibiting protein

The functional elucidation of small unknown phage proteins (‘ORFans’) presents itself as one of the major challenges of bacteriophage molecular biology. In this work, we mined the Pseudomonas aeruginosa infecting phage LUZ24 proteome for antibacterial and antibiofilm proteins against its host. Subsequently, their putative host target was identified. In one example, we observed an interaction between LUZ24 gp4 and the host transcriptional regulator MvaT. The polymerization of MvaT across AT-rich DNA strands permits gene silencing of foreign DNA, thereby limiting any potentially adverse effects of such DNA. Gel shift assays proved the inhibitory effect of LUZ24 gp4 on MvaT DNA binding activity. Therefore, we termed this gene product as Mip, the MvaT inhibiting protein. We hypothesize Mip prevents the AT-rich LUZ24 DNA from being physically blocked by MvaT oligomers right after its injection in the host cell, thereby allowing phage transcription and thus completion of the phage infection cycle