Aim. Staphylococcus aureus is the first causative agent of bone and joints infections (BJI). It causes difficult-to-treat infections because of its ability to form biofilms, and to be internalized and persist inside osteoblastic cells. Recently, phage therapy has emerged as a promising therapy to improve the management of chronic BJI. In the present study, we evaluated the efficacy of an assembly of three bacteriophages previously used in a clinical case report (Ferry, 2018) against S. aureus in in vitro models of biofilm and intracellular osteoblast infection. Methods. Using HG001 S. aureus, the bactericidal activities of the assembly of the three bacteriophages (Pherecydes Pharma) used alone or in association with vancomycin or rifampicin were compared by quantifying the number of viable bacteria in mature biofilms and infected osteoblasts after 24h of exposure. Results. The activity of bacteriophages against biofilm-embedded S. aureus was dose-dependent. Synergistic effects were observed when bacteriophages were combined to antibiotics at the lowest concentrations, with no significant bactericidal activity in monotherapy. In the human osteoblast infection model, we were able to show that phage penetration into osteoblasts was only possible when the cells were infected, suggesting a S. aureus dependent Trojan horse mechanism. The intracellular inoculum in osteoblasts treated with bacteriophages or vancomycin was significantly higher than in cells treated with lysostaphin, used as control condition of rapid killing of bacteria released in the extracellular media after death of infected cells and absence of intracellular activity. These results suggest that bacteriophages are probably both i) inactive in the intracellular compartment and ii) unable to kill all bacteria released after cell lysis into the extracellular medium fast enough to prevent them from reinfecting other osteoblasts. Conversely, the intracellular inoculum recovered from cells treated with vancomycin+bacteriophages was significantly lower than the one inside cells treated with vancomycin or bacteriophages alone, suggesting that this combination allowed a better control of released bacteria in the extracellular media. Finally, bacteriophages did not increase the activity of rifampicin in this model. Conclusion. In conclusion, we showed that the bacteriophages tested were highly active against S. aureus in mature biofilm but had no activity against bacteria internalized in osteoblasts. Additional studies using animal models of BJI and well-conducted clinical trials are needed to further evaluate phage therapy and its positioning in the management of these infections

Aim. Bacteriophages, viruses specific of bacteria, are receiving substantial attention as alternative antibacterial agents to treat bacteria frequently multi-resistant to antibiotics and/or able to form biofilms, such as staphylococci. The latter are responsible for very difficult to treat bone and joint infections (BJIs). In this context, our consortium aims to develop a production of therapeutic phages in accordance with the will of ANSM (French National Agency for the Safety of Medicines and Health Products) to encourage the development of a national academic platform for phage therapy. We report the isolation and characterization of new anti-Staphylococcus phages as well as the evaluation of their activity on a collection of clinical strains of S. aureus (SA) and coagulase-negative staphylococci (CNS) in order to assess their therapeutic potential. Method. Seventeen phages were isolated from wastewater samples. Their identification was obtained by Illumina whole genome sequencing. To evaluate their spectrum of activity, 30 genetically characterized SA strains representative of the main genetic backgrounds as well as 32 strains belonging to 7 CNS species responsible for BJIs were included. The spot test technique, based on the determination of the Efficiency Of Plating ratio, was used (EOP, ratio between the phage titer obtained on a tested strain/titer on a reference strain, close to 1 if high sensitivity to the phage). Results. All isolated phages belonged to the Myoviridae family: 14/17 and 3/17 to the Kayvirus and Silviavirus genera respectively. Silviavirus phages were more active on SA strains (EOP>0.001 for 73–90% of strains) than Kayvirus phages (EOP>0.001 for 13–70% of strains, except for V1SA21: 80%). In total, 83% of strains were susceptible to the phage with the broadest spectrum in each genus, their combination representing a promising opportunity to prevent the emergence of resistance. Kayvirus phages had polyvalent activity on several CNS species (maximum 47% of tested strains), mainly S. lugdunensis, S. capitis and S. caprae, whereas Silviavirus phages were only active on 6–12% of the tested strains. Conclusions. We report the characterization of a large collection of novel phages with complementary spectra against a collection of SA and CNS strains. Further work is currently focused on i) the isolation of anti-S. epidermidis phages, bacterial species against which the present collection of phages was insufficiently active, while it is a major pathogen in this context, ii) the development of production and purification protocols in order to meet the requirements of ANSM for human use

Aim

Many bone and joint infections, in spite of appropriated antibiotics therapy and surgery, lead to a therapeutic dead end. We are then faced with a chronic infection with or without continuous antibiotic treatment, with daily local care, and an exhorbitant economic and social cost. Pami the incriminated factors: the presence of foreign implant material, the poor diffusion of antibiotics at the infectious site, the presence of biofilm.

The bacteriophages, biological drug, natural environmental viruses possess the properties to meet these difficulties: well diffusion to the infectious focus with possibilities of local use, destruction of the biofilm allowing a release of the bacteria and a synergistic effect with the antibiotics, antibiofilm effect for the restoration of osteoblastosis.

Aim. Phage therapy has attracted attention as a promising alternative treatment option for biofilm infections. To establish a successful phage therapy, a comprehensive stock of different phages covering a broad bacterial spectrum is crucial. We screened human and environmental sources for presence of lytic phages against selected bacteria. Methods. Saliva collected from 10 volunteers and 500 ml of sewage water were screened for the presence of lytic phages active against 20 clinical strains of Staphylococcus aureus and 10 of Escherichia coli, both isolated from patients with prosthetic joint infection. Laboratory strains of methicillin-resistant S. aureus (MRSA)*1 and E. coli*2 were also tested. Screening was performed plaque-assay to detect phages for different strains. Isolated plaques were collected and phages were enriched to determine their activity against their bacterial host strains. The activity of bacteriophages against adherent E. coli and MRSA was evaluated by crystal violet, staining bacterial biofilms grown on glass beads. Results. Six bacteriophages specific for MRSA were isolated from saliva. Bacteriophages for E. coli strains were isolated from sewage water (n=3) and saliva (n=1). All bacteriophages tested against biofilms of their bacterial host showed a reduction of the total biomass (ranging from 19% to 84%). Conclusions. Both sewage and saliva samples provided bacteriophages specific against selected bacterial strains. 24h phage treatment of E. coli and S. aureus biofilms lead to a reduction but not to a complete eradication of biofilm

Background. Bacteriophages are natural viruses of interest in the field of PJI. A paper previously reported the PhagoDAIR procedure (use of phages during DAIR) in three patients with PJI for whom explantation was not desirable. As the need to isolate the pathogen before surgery to perform phage susceptibility testing is a strong hindrance for the development of this procedure, we developed post-operative phage injections using ultrasound, in patients infected with S. aureus and/or P. aeruginosa who were eligible for the PhagoDAIR procedure, but for whom phages were not available at the time of surgery. Materials/Methods. We performed a single center, exploratory, prospective cohort study including patients with knee PJI who received phage therapy with ultrasound after performance of a DAIR or a partial prosthesis exchange. All patients had PJI requiring conservative surgery and suppressive antimicrobial therapy (SAT) as salvage procedure. Each case was discussed in multidisciplinary meetings in agreement with French health authority, based on the clinical presentation, and the phage susceptibility testing. The cocktail of highly concentrate active phages (5 mL; about 10. e. 9 PFU/mL) was extemporaneous prepared and administered three times directly into the joint using sonography (1 injection per week during 3 weeks) during the postoperative period, before switching antibiotics to SAT. Results. Seven patients received phages under sonography after the DAIR, and one after a partial exchange (mean age 71 years). All had resection prosthesis or constrained knee prosthesis. Among these seven patients, three were infected with S. aureus (including one MRSA), two were infected with P. aeruginosa (one was a multidrug-resistant isolate), one was infected with both S. aureus and P. aeruginosa and the last one was infected with MRSA, S. epidermidis and Corynebacterium spp.. All patients received a cocktail of active phages provided by Pherecydes Pharma targeting S. aureus or P. aeruginosa. No adverse event was recorded during or after the local injections. All patients were switched to SAT after a primary postoperative antimicrobial therapy of three months. Under SAT, the patient with S. epidermidis co-infection developed a relapse due to the S. epidermidis. With a mean follow-up of 13 months after surgery (from 9 to 24 months), the outcome was favorable for all patients without any sign of infection; none of them had abnormal pain, joint effusion or loosening. Conclusions. Postoperative administration of phages using sonography is a potentially useful procedure in patients with complex PJI for whom a conservative approach is desirable

The number of arthroplasties being undertaken is expected to grow year on year, and periprosthetic joint infections will be an increasing socioeconomic burden. The challenge to prevent and eradicate these infections has resulted in the emergence of several new strategies, which are discussed in this review.

Cite this article: Bone Joint J 2015;97-B:1162–9.