Rapid Isolation, Propagation, and Online Analysis of a Small Number of Therapeutic Staphylococcal Bacteriophages from...

A method for the fast isolation, propagation, and characterization of very low count bacteriophages active against pathogenic bacterial strains is described in this study. Bacteriophages with a count of 10² phage particles were dynamically adhered from the maximum 10 mL blood plasma sample onto the nanostructured part of the fused silica capillary. One-step propagation of phage particles of genus Kayvirus inside the etched capillary on 10⁴Staphylococcus aureus host cells increased their number to 6 × 10⁴ phage particles. Phage particles were concentrated online and separated by capillary electrophoretic methods. No phage replication occurred when the phage-resistant S. aureus or Escherichia coli cells were used. Two-step phage propagation in the capillary allowed an increase in the total virion count to up to 6 × 10⁵ phage particles and subsequent off-line matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of the phage zone collected after capillary electrophoresis. Relative standard deviations of the phage peak area were at most 2.3%. We expect that the method of isolating bacteriophages from blood plasma and their simultaneous identification will facilitate clinical studies of phage preparations and contribute to pharmacokinetics studies during phage therapy. This approach is also suitable for capturing and enriching new phages from the environment when a susceptible indicator strain is available.