Phage therapy might be a useful approach for the treatment of nosocomial infections; however, only few lytic phages suitable for this application are available for the opportunistic pathogen, Staphylococcus epidermidis. In the current study, we developed an efficient method to isolate bacteriophages present within the human skin microbiome, by using niche-specific S. epidermidis as the host for phage propagation. S.epidermidis was identified on the forehead of 92% of human subjects tested. These isolates were then used to propagate phages present in the same skin sample. Plaques were observable on bacterial lawns in 46% of the cases where S. epidermidis was isolated. Eight phage genomes were genetically characterized, including the previously described phage 456. Six phage sequences were unique, and spanned each of the major staphylococcal phage families; Siphoviridae (n = 3), Podoviridae (n = 1), and Myoviridae (n = 2). One of the myoviruses (vB_SepM_BE06) was identified on the skin of three different humans. Comparative analysis identified novel genes including a putative N-acetylmuramoyl-L-alanine amidase gene. The host-range of each unique phage was characterized using a panel of diverse staphylococcal strains (n = 78). None of the newly isolated phages infected more than 52% of the S. epidermidis strains tested (n = 44), and non-S. epidermidis strains where rarely infected, highlighting the narrow host-range of the phages. One of the phages (vB_SepM_BE04) was capable of killing staphylococcal cells within biofilms formed on polyurethane catheters. Uncovering a richer diversity of available phages will likely improve our understanding of S. epidermidis-phage interactions, which will be important for future therapy.
Pseudomonas aeruginosa
is a major pathogen in burn wound infections. We present one of the first reports of small-colony variant (SCV) emergence of
P. aeruginosa
, taken from a patient under aminoglycosides for a persistent burn wound infection. We confirm the causative role of a single
ispA
mutation in SCV emergence and increased aminoglycoside resistance. IspA is involved in the synthesis of ubiquinone, providing a possible link between electron transport and SCV formation in
P. aeruginosa
.
Pseudomonas aeruginosa
is a Gram-negative pathogen causing infections in acutely burned patients. The precise mechanisms required for the establishment of infection in the burn setting, and adaptive traits underpinning prolonged outbreaks are not known.
Background
Treatment failure in pneumococcal meningitis due to antibiotic resistance is an increasing clinical challenge and alternatives to antibiotics warrant investigation. Phage-derived endolysins efficiently kill gram-positive bacteria including multi-drug resistant strains, making them attractive therapeutic candidates. The current study assessed the therapeutic potential of the novel endolysin PlyAZ3aT in an infant rat model of ceftriaxone-resistant pneumococcal meningitis.
Methods
Efficacy of PlyAZ3aT was assessed in a randomized, blinded and controlled experimental study in infant Wistar rats. Meningitis was induced by intracisternal infection with 5 x 107 CFU/ml of a ceftriaxone-resistant clinical strain of S. pneumoniae, serotype 19A. Seventeen hours post infection (hpi), animals were randomized into 3 treatment groups and received either (i) placebo (phosphate buffered saline [PBS], n = 8), (ii) 50 mg/kg vancomycin (n = 10) or (iii) 400 mg/kg PlyAZ3aT (n = 8) via intraperitoneal injection. Treatments were repeated after 12 h. Survival at 42 hpi was the primary outcome; bacterial loads in cerebrospinal fluid (CSF) and blood were secondary outcomes. Additionally, pharmacokinetics of PlyAZ3aT in serum and CSF was assessed.
Results
PlyAZ3aT did not improve survival compared to PBS, while survival for vancomycin treated animals was 70% which is a significant improvement when compared to PBS or PlyAZ3aT (p<0.05 each). PlyAZ3aT was not able to control the infection, reflected by the inability to reduce bacterial loads in the CSF, whereas Vancomycin sterilized the CSF and within 25 h. Pharmacokinetic studies indicated that PlyAZ3aT did not cross the blood brain barrier (BBB). In support, PlyAZ3aT showed a peak concentration of 785 μg/ml in serum 2 h after intraperitoneal injection but could not be detected in CSF.
Conclusion
In experimental pneumococcal meningitis, PlyAZ3aT failed to cure the infection due to an inability to reach the CSF. Optimization of the galenic formulation e.g. using liposomes might enable crossing of the BBB and improve treatment efficacy.
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