Proof-of-Concept Standardized Approach Using a Single-Disk Method Analogous to Antibiotic Disk Diffusion Assays for Routine Phage Susceptibility Testing in Diagnostic Laboratories

Application of bacteriophages is increasingly being implemented in clinical therapies. Prior susceptibility testing should be regarded as mandatory, but standards are lacking. The objective of this research was to develop a highly standardized methodology to facilitate phage susceptibility testing (PST) in clinical microbiology routine laboratories. Therefore, EUCAST methods established for single disk-based antibiotic susceptibility testing (AST) were adapted. In a first step, basic parameters were evaluated using well-studied Escherichia phage T4-Escherichia coli combinations. In addition, test results were compared to those from conventional spot test and efficiency of plating (EOP) approaches. In a second step, the applicability of the methodology and the most promising test parameters were demonstrated for five other frequently isolated clinical bacterial species and their corresponding phages. At present, the method predominantly leads to qualitative rather than quantitative results. This disk-based approach provides a standardized, easy-to-handle, reproducible and reliable PST protocol by relying on well-established routine procedures in diagnostic laboratories.IMPORTANCE Application of bacteriophages in clinical therapies is attractive due to increasing rates of isolation of multidrug-resistant bacteria worldwide. As the phage effect is highly specific, prior susceptibility testing of target bacteria is mandatory. Of note, established standards are lacking. In this research, we adapted the single-disk method for antibiotic susceptibility testing to phage susceptibility testing (PST) in order to provide a standardized, easy-to-handle, reproducible, and reliable PST protocol for application in diagnostic routine laboratories. Application of bacteriophages in clinical therapies is attractive due to increasing rates of isolation of multidrug-resistant bacteria worldwide. As the phage effect is highly specific, prior susceptibility testing of target bacteria is mandatory.